Cyclosporin a analogs

ABSTRACT

The present invention relates to novel cyclosporin analogs, processes for preparing them, pharmaceutical compositions containing them, and methods for using these analogs and the compositions containing them for the treatment of medical conditions, including but not limited to ocular conditions such as dry eye.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional PatentApplication Ser. No. 61/654,388, filed on Jun. 1, 2012, the disclosureof which is incorporated herein by reference.

FIELD OF THE INVENTION

The present invention relates to novel cyclosporin analogs, processesfor preparing them, pharmaceutical compositions containing them, andmethods for using these analogs and the compositions containing them forthe treatment of medical conditions, including but not limited to ocularconditions such as dry eye.

BACKGROUND OF THE INVENTION

Cyclosporins are a class of poly-N-methylated cyclic undecapeptides. Thefirst cyclosporin to be isolated was cyclosporin A (CAS Registry Number:59865-13-3), a naturally occurring fungal metabolite having thefollowing structure:

As shown by the structure above, Cyclosporin A consists of 11 aminoacids and can be further represented as follows:

where:

MeBmt is (4R)-4-[(E)-2-butenyl]-4-methyl-L-threonine;

αAbu is L-α-aminobutyric acid;

Sar is sarcosine;

MeLeu is N-methyl-L-leucine;

Val is L-valine;

Ala is L-alanine;

DAla is D-alanine; and

MeVal is N-methyl-L-valine.

The numbers 1-11 are used to designate each of the eleven amino acids.Thus, MeBMT is the amino acid at position 1; sarcosine, the amino acidat position 3. In certain instances, the description herein may refer tothe amino acid side chain at any one of positions 1-11. The carbon towhich the amino acid side chain is attached is referred to as the alpha(α) carbon.

Cyclosporin A is best known for its immunosuppressive properties and iscommonly prescribed for use in patients that have undergone bone marrowor organ transplantation. The present invention relates to the discoveryof water-soluble analogs of cyclosporin A that are potent inhibitors ofcyclophilin.

SUMMARY OF THE INVENTION

Accordingly, the present invention describes compounds having Formula I:

wherein:R¹ is —H, —C₁₋₆alkyl, —OC₁₋₆alkyl, —CH₂F, —CH₂OCH₃, —SC₁₋₆alkyl, —CH₃,—CH₂CH₃, —SCH(CH₃)₂, —CH₂OH, —SCH₃, —OCH₃, —R¹³R¹⁴,

R² is —CH₃, —CH₂CH₃, or —CH₂CH₂CH₃;

R³ is —CH₂CH(CH₃)₂, —CH(CH₃)₂, —CH₂C(CH₃)₂(OH), —CH(CH₃)(CH₂CH₃), or—CH₂CH(R⁷)(CH₂CH₃);

R⁴ is —CH₃, or —CH₂OH;

R⁵ is —R⁸(CH₂)_(n)(C═O)_(m)—

R⁶ is —CH₂CH₃, —CH(CH₃)(OH), —CH(CH₃)₂, or —CH₂CH₂CH₃;

R⁷ is OC₁₋₅ alkyl;

R⁸ is O, S, CH₂O, CH₂S, or CH₂;

R⁹ is —H, —C₁₋₅ alkyl, —C₂₋₄fluoroalkyl, —C₁₋₅ alkyl-heterocycle,cyanoalkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or R⁹ taken togetherwith R¹¹, R¹⁰, and the N to which R⁹ and R¹⁰ are attached forms aheterocycle;R¹⁰ is —H, —C₁₋₅ alkyl, —C₂₋₄fluoroalkyl, —C₁₋₅ alkyl-heterocycle,cyanoalkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or R¹⁰ takentogether with R¹¹, R⁹, and the N to which R⁹ and R¹⁰ are attached formsa heterocycle;R¹¹ is O, NR¹², S(O)_(q), CF₂, C₁₋₅alkylene, CH(OC₁₋₆alkyl), divalentC₃₋₈cycloalkyl, divalent heterocycle, carbonyl, or taken together withR⁹, R¹⁰, and the N to which R⁹ and R¹⁰ are attached forms a heterocycle;R¹² is H, CH₃, or C₁₋₅ alkyl;

R¹³ is O, S, CH₂O, CH₂S, CH₂SO, or CH₂SO₂;

R¹⁴ is H, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, C₃₋₈cycloalkyl,CH₂CH₂N(CH₂CH₃)₂, CH₂CH₂NH(CH₂CH₃), heterocycle, or aryl;n=0, 1, 2, 3, or 4;m=0 or 1;p=0 or 1; andq=0, 1, or 2;wherein R¹⁴ is optionally substituted with one or more groupsindependently selected from the group consisting of H, C₁₋₆alkyl,halogen, hydroxyl, ester, sulfonamide, ketone, aldehyde, cycloalkyl,heterocycle, aryl, amine, heterocycle, amide, and guanidinyl;wherein the heterocycle comprising R⁹, R¹⁰, R¹¹, and the N to which R⁹and R¹⁰ are attached is monocyclic or polycyclic;wherein “- - -” is a single bond or is absent; andwith the provisos thatwhen R⁸ is O, S, CH₂O, or CH₂S then n is not 0 or 1;when p=0 then R¹¹ and “- - -” are absent; andwhen R¹¹ and “- - -” are absent then R⁹ is not directly linked to R¹⁰.

In another embodiment, the invention provides for a compound havingFormula I, wherein:

R¹ is —H, —C₁₋₆alkyl, —OC₁₋₆alkyl, —CH₂F, —CH₂OCH₃, —SC₁₋₆alkyl, —CH₃,—CH₂CH₃, —SCH(CH₃)₂, —CH₂OH, —SCH₃, —OCH₃, —R¹³R¹⁴,

R² is —CH₃, —CH₂CH₃, or —CH₂CH₂CH₃;

R³ is —CH₂CH(CH₃)₂, —CH(CH₃)₂, —CH₂C(CH₃)₂(OH), —CH(CH₃)(CH₂CH₃), or—CH₂CH(R⁷)(CH₂CH₃);

R⁴ is —CH₃, or —CH₂OH;

R⁵ is —R⁸(CH₂)_(n)(C═O)_(m)—

R⁶ is —CH₂CH₃, —CH(CH₃)(OH), —CH(CH₃)₂, or —CH₂CH₂CH₃;

R⁷ is OC₁₋₅ alkyl;

R⁸ is O, S, CH₂O, CH₂S, or CH₂;

R⁹ is —H, —C₁₋₅ alkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or taken togetherwith R¹¹, R¹⁰, and the N to which R⁹ and R¹⁰ are attached forms aheterocycle;R¹⁰ is —H, —C₁₋₅ alkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or taken togetherwith R¹¹, R⁹, and the N to which R⁹ and R¹⁰ are attached forms aheterocycle;R¹¹ is O, NR¹², S(O)_(q), C₁₋₅alkylene, divalent C₃₋₈cycloalkyl,divalent heterocycle, carbonyl, or taken together with R⁹, R¹⁰, and theN to which R⁹ and R¹⁰ are attached forms a heterocycle;R¹² is H, CH₃, or C₁₋₅ alkyl;

R¹³ is O, S, CH₂O, CH₂S, CH₂SO, or CH₂SO₂;

R¹⁴ is H, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, C₃₋₈cycloalkyl,CH₂CH₂N(CH₂CH₃)₂, CH₂CH₂NH(CH₂CH₃), heterocycle, or aryl;n=0, 1, 2, 3, or 4;m=0 or 1;p=0 or 1; andq=0, 1, or 2;wherein R¹⁴ is optionally substituted with one or more groupsindependently selected from the group consisting of H, C₁₋₆alkyl,halogen, hydroxyl, ester, sulfonamide, ketone, aldehyde, cycloalkyl,heterocycle, aryl, amine, heterocycle, amide, and guanidinyl;wherein the heterocycle comprising R⁹, R¹⁰, R¹¹, and the N to which R⁹and R¹⁰ are attached is monocyclic or polycyclic;wherein “- - -” is a single bond or is absent; andwith the provisos thatwhen R⁸ is O, S, CH₂O, or CH₂S then n is not 0 or 1;when p=0 then R¹¹ and “- - -” are absent; andwhen R¹¹ and “- - -” are absent then R⁹ is not directly linked to R¹⁰.

A compound having Formula I according to any of the embodiments setforth above may further include the proviso that when R² is —CH₃, and R³is —CH₂CH(CH₃)₂, and R⁴ is —CH₃, and

R⁶ is —CH₂CH₃, then the group

-   -   is not —CH₂CH₂CH₂(C═O)N(CH₂CH₃)₂, —CH₂CH₂CH₂(C═O)N(CH₃)₂,        —CH₂CH₂CH₂(C═O)NH₂, —CH₂CH₂CH₂CH₂CH₂NH(C═O)CH₃,        —CH₂CH₂CH₂CH₂NH(C═O)CH₃, —CH₂CH₂CH₂CH₂CH₂NH(C═O)CH₂CH₃,        —CH₂CH₂CH₂CH₂CH₂NH(C═O)CH₂CH₂ CH₃, —CH₂CH₂CH₂CH₂NH(C═O)CH₃,        —CH₂NHCH₂COOH, or —CH₂NH(CH₂)₅COOH.

Additionally or alternatively, a compound according to any of theembodiments set forth above may further include the provisos that whenm=1, and R² is —CH₃, and R³ is —CH₂CH(CH₃)₂, and R⁴ is —CH₃, and R⁶ is—CH₂CH₃ then neither R⁹ nor R¹⁰ is —H or —C₁-C₅alkyl, and R⁹, R¹⁰, R¹¹,and the N to which R⁹ and R¹⁰ are attached taken together do not formmorpholinyl.

A compound having Formula I may further include the provisos that whenm=1 then neither R⁹ nor R¹⁰ is —H or —C₁-C₅alkyl, and R⁹, R¹⁰, R¹¹, andthe N to which R⁹ and R¹⁰ are attached taken together do not formmorpholinyl.

In one embodiment the invention provides a compound having Formula I,wherein R¹ is not hydrogen (H).

In another embodiment the invention provides a compound having FormulaI, wherein when R¹ is H, then m=0.

In another embodiment the invention provides a compound having FormulaI, wherein when R¹ is H, then m=0 and R⁹, R¹⁰, R¹¹, and the N to whichR⁹ and R¹⁰ are attached together form a heterocycle.

In another embodiment the invention provides a compound having FormulaI, wherein R⁶ is —CH₂CH₃.

In another embodiment the invention provides a compound having FormulaI, wherein R⁶ is —CH(CH₃)(OH).

In another embodiment the invention provides a compound having FormulaI, wherein R⁶ is —CH(CH₃)₂.

In another embodiment the invention provides a compound having FormulaI, wherein R⁶ is —CH₂CH₂CH₃.

In another embodiment the invention provides a compound having FormulaI, wherein n=0, 1, 2, 3, or 4 and m=0.

In another embodiment the invention provides a compound having FormulaI, wherein n=0, 1, 2, 3, or 4 and m=1.

In another embodiment the invention provides a compound having FormulaI, wherein n=0, m=0, and p=1.

In another embodiment the invention provides a compound having FormulaI, wherein n=1, m=1, and p=1.

In another embodiment the invention provides a compound having FormulaI, wherein n=1, m=1, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein n=3, m=0, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein n=3, m=0, and p=1.

In another embodiment the invention provides a compound having FormulaI, wherein n=2, m=0, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R⁸ is CH₂O, n=2, and m=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₂CH₃, R¹¹ is O, n=0, and p=1.

In another embodiment the invention provides a compound having FormulaI, wherein R² is —CH₃ or —CH₂CH₃; R³ is —CH₂CH(CH₃)₂, —CH(CH₃)₂, or—CH₂C(CH₃)₂(OH); R⁴ is —CH₃; R⁶ is —CH₂CH₃; and R⁸ is CH₂.

In another embodiment the invention provides a compound having FormulaI, wherein R² is —CH₃; R³ is —CH₂CH(CH₃)₂; R⁴ is —CH₃; R⁶ is —CH₂CH₃;and R⁸ is CH₂.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, —CH₂CH₃, or —SCH₃; R² is —CH₃; R³ is—CH₂CH(CH₃)₂; R⁴ is —CH₃; R⁶ is —CH₂CH₃; and R⁸ is CH₂.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —SCH₃, R¹¹ is O, n=0, m=0 and p=1, and wherein R⁹, R¹⁰,R¹¹, and the N to which R⁹ and R¹⁰ are attached taken together form aheterocycle.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁶ is—CH₂CH₃, R⁸ is CH₂, n=0, 1, 2, 3, or 4, m=0, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁶ is—CH₂CH₃, R⁸ is CH₂, n=0, 1, 2, 3, or 4, m=0, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂CH₂CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, n=2, m=0, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —R¹³R¹⁴, R¹¹ is O, R¹³ is CH₂S, R¹⁴ is—CH₂CH₂N(CH₂CH₃)₂, n=0, m=0 and p=1, and wherein R⁹, R¹⁰, R¹¹, and the Nto which R⁹ and R¹⁰ are attached taken together form a heterocycle.

In another embodiment the invention provides a compound having FormulaI, wherein R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁶ is —CH₂CH₃,and R⁸ is CH₂.

In another embodiment the invention provides a compound having FormulaI, wherein R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁶ is —CH₂CH₃,and R⁸ is CH₂O.

In another embodiment the invention provides a compound having FormulaI, wherein R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁶ is —CH(CH₃)₂,and R⁸ is CH₂.

In one embodiment the invention provides a compound having Formula I,wherein p=1 and R⁹, R¹⁰, R¹¹ and the N to which R⁹ and R¹⁰ are attachedtaken together form a heterocycle.

In another embodiment the invention provides a compound having FormulaI, wherein p=0 and R⁹ is not directly linked to R¹⁰.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃ and R⁶ is —CH₂CH₃.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R⁶ is —CH₂CH₃ or —CH(CH₃)₂, R⁸ is CH₂, R¹¹ is O,n=0, 1, 2, 3, or 4, p=1, and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰are attached taken together form morpholine.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R⁶ is —CH₂CH₃ or —CH(CH₃)₂, R⁸ is CH₂, R¹¹ isNR¹², R¹² is CH₃, n=0, 1, 2, 3, or 4, p=1, and R⁹, R¹⁰, R¹¹, and the Nto which R⁹ and R¹⁰ are attached taken together form N-methylpiperazine.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R⁶ is —CH₂CH₃ or —CH(CH₃)₂, R⁸ is CH₂, R¹¹ isNR¹², R¹² is H, n=0, 1, 2, 3, or 4, p=1, and R⁹, R¹⁰, R¹¹, and the N towhich R⁹ and R¹⁰ are attached taken together form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃ or —CH₂CH₃, R³ is —CH₂CH(CH₃)₂ or—CH(CH₃)₂, R⁴ is —CH₃, and R⁶ is —CH₂CH₃, R⁸ is CH₂, R¹¹ is O or NR¹²,R¹² is CH₃, m=1, n=2, and p=1, and wherein R⁹, R¹⁰, R¹¹ and the N towhich R⁹ and R¹⁰ are attached taken together form a heterocycle

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —H, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁶ is—CH₂CH₃, R⁸ is CH₂, R¹¹ is O, n=3, m=0, and p=1, and wherein R⁹, R¹⁰,R¹¹ and the N to which R⁹ and R¹⁰ are attached taken together form aheterocycle.

In another embodiment the invention provides a compound having FormulaI, wherein m=1, p=1, and wherein R⁹, R¹⁰, R¹¹ and the N to which R⁹ andR¹⁰ are attached taken together form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R⁹ is —H, R¹⁰ is

and m=1.

In another embodiment the invention provides a compound having FormulaI, wherein R⁹ is —H, R¹⁰ is

and m=1.

In another embodiment the invention provides a compound having FormulaI, wherein R⁹ is —H, R¹⁰ is

and m=1.

In another embodiment the invention provides a compound having FormulaI, wherein R⁹ is —CH₂CH₃, R¹⁰ is —CH₂CH₃, m=1, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, n=1, m=1, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂CH₂CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, n=2, m=0, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂OCH₂CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂O, n=2, m=0, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₂CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵is —CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, R¹¹ is O, n=0, m=0, and p=1, andwherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached takentogether form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —SCH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵is —CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, R¹¹ is O, n=0, m=0, and p=1, andwherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached takentogether form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂CH₂CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, n=2, m=0, and p=0.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —R¹³R¹⁴, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵is —CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, R¹¹ is O, R¹³ is CH₂S, R¹⁴ is—CH₂CH₂N(CH₂CH₃)₂, n=0, m=0 and p=1, and wherein R⁹, R¹⁰, R¹¹, and the Nto which R⁹ and R¹⁰ are attached taken together form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂—, R⁶ is —CH(CH₃)₂, R⁸ is CH₂, R¹¹ is O, n=0, m=0, and p=1, andwherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached takentogether form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —OCH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵is —CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, R¹¹ is O, n=0, m=0, and p=1, andwherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached takentogether form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂CH₂CH₂CH₂—, R⁶ is —CH(CH₃)₂, R⁸ is CH₂, R¹¹ is O, n=3, m=0, and p=1,and wherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —H, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂CH₂CH₂CH₂—, R⁶ is —CH(CH₃)₂, R⁸ is CH₂, R¹¹ is O, n=3, m=0, and p=1,and wherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form

wherein “- - -” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, R¹¹ is O, n=0, m=0, and p=1, andwherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached takentogether form

wherein “*” represents the point of attachment to R⁵.

In another embodiment the invention provides a compound having FormulaI, wherein R¹ is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is—CH₂CH₂CH₂CH₂—, R⁶ is —CH₂CH₃, R⁸ is CH₂, R¹¹ is O, n=3, m=0, and p=1,and wherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form

wherein “*” represents the point of attachment to R⁵.

In some embodiments, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, and R⁶is —CH₂CH₃.

In some embodiments R⁵ is —R⁸(CH₂)_(n)(C═O)_(m)—, which is attached tothe nitrogen bearing the R⁹ and R¹⁰ groups through the (C═O), R⁸ is CH₂,n=0, 1, 2, or 3, and m=0. Thus, in some embodiments R⁵ is —CH₂,—CH₂CH₂—, —CH₂CH₂CH₂—, or —CH₂CH₂CH₂CH₂—. In some embodiments R⁸ is CH₂,n=1, and m=1. Accordingly, in some embodiments R⁵ is —CH₂CH₂(C═O)—, forexample. In some embodiments R⁸ is CH₂O, n=2, and m=0. Accordingly, insome embodiments R⁵ is —CH₂OCH₂CH₂—, for example.

Non-limiting examples of R¹¹ include O, NCH₃, NH, SO₂, S, CH₂, CF₂, andCH(OCH₃).

Non-limiting examples of a C₁₋₆alkyl-heterocycle include —CH₂Pyrid-2-yl,—CH₂Pyrid-3-yl, —CH₂Pyrimidin-2-yl, —CH₂Pyrazin-2-yl,—CH₂-3-Me-Imidazol-4-yl, —CH₂-2-Me-Pyrazol-3-yl, —CH₂Pyrid-4-yl,—CH₂-1-Me-Pyrazol-4-yl, and —CH₂-1-Me-3-CF₃-Pyrazol-5-yl.

A non-limiting example of a cyanoalkyl includes —CH₂CH(CH₃)CN.

In some embodiments, R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form a heterocycle. In some embodiments theheterocycle is a non-aromatic heterocycle. In other embodiments theheterocycle is an aromatic heterocycle.

Non-limiting examples of compounds having Formula I include those inwhich m=0 and R¹ is not H, and/or in which p=1 and R⁹, R¹⁰, R¹¹, and theN to which R⁹ and R¹⁰ are attached together form a heterocycle. Othernon-limiting examples of compounds having Formula I include thosewherein R¹ is —CH₃, —SCH₃, —OCH₃, or —CH₂OH; R⁸ is CH₂; n=0, 1, 2, 3, or4; m=0; p=1, and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form a heterocycle. Additional examples ofcompounds include those wherein n=0, 1, 2, 3, or 4, m=0, p=0 or 1, andR¹ is —C₁₋₆alkyl, —OC₁₋₆alkyl, or —SC₁₋₆alkyl.

In some embodiments, the R⁹ and R¹⁰ heterocycle, or the heterocycleformed from R⁹, R¹⁰, and the N to which R⁹ and R¹⁰ are attached isindependently selected from the group consisting of piperidyl,pyrrolidinyl, piperazinyl, morpholinyl, thiomorpholinyl, thiazolidinyl,1,4-dioxanyl, tetrahydrofuranyl, tetrahydrothiophenyl, lactam, lactone,pyridyl, pyrazinyl, furanyl, thienyl, pyrimidinyl, pyridone (includingN-substituted pyridones), isoxazolyl, isothiazolyl, oxazolyl, thiazolyl,pyrazolyl, furazanyl, pyrrolyl, pyrazolyl, triazolyl,1,2,4-thiadiazolyl, pyrazinyl, pyridazinyl, quinoxalinyl, phthalazinyl,oxindolyl, imidazo[1,2-a]pyridinyl, imidazo[2,1-b]thiazolyl,benzofurazanyl, indolyl, azaindolyl, benzimidazolyl, benzothienyl,quinolinyl, imidazolyl, thienopyridyl, quinazolinyl, thienopyrimidyl,pyrrolopyridyl, imidazopyridyl, isoquinolinyl, benzoazaindolyl,1,2,4-triazinyl, benzothiazolyl,

wherein “*” indicates the point of attachment to R⁵.

The heterocycle formed by R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰are attached taken together may be optionally substituted by an alkyl,halogen, or haloalkyl. One example of a haloalkyl is —CF₃. In someembodiments one or more hydrogen atoms on the heterocycle is replacedwith fluorine (F). In specific embodiments, the fluorinated heterocycleis a saturated, non-aromatic heterocycle. In a more specific embodimentthe fluorinated heterocycle is

wherein “*” represents the point of attachment to R⁵.

In some embodiments, the compound having Formula I is any one of thoselisted in Tables 1-27. Accordingly, non-limiting examples of a compoundaccording to the present invention include any of the compounds listedin Tables 1-27. The present invention also provides for a pharmaceuticalcomposition comprising or consisting of a therapeutically effectiveamount of a compound having Formula I and a pharmaceutically acceptableexcipient.

For example, one embodiment is a compound having Formula I, whereinR¹═—CH₃, R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, R¹¹ isO, n=0, m=0, p=1, and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:

wherein “*” represents the point of attachment to R⁵ (Compound F).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, R⁹═—H, R¹⁰ is

n=1, m=1, and p=0 (Compound W).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, n=1, m=1, p=1,q=2, R¹¹ is S(O)_(q), and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰are attached taken together form:

wherein “*” represents the point of attachment to R⁵ (Compound Z).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, n=1, m=1, p=1,and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached takentogether form:

wherein “*” represents the point of attachment to R⁵ (Compound ZZ).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, n=0, m=0, p=1,and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached takentogether form:

wherein “*” represents the point of attachment to R⁵ (Compound O).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, R¹¹ is O, n=2,m=0, p=1, R¹¹ is O, and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:

wherein “*” represents the point of attachment to R⁵ (Compound KG).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, n=0, m=0, p=1,R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:

wherein “*” represents the point of attachment to R⁵ (Compound EK).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, n=0, m=0, p=1,R¹¹ is CF₂, and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:

wherein “*” represents the point of attachment to R⁵ (Compound EL).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, n=0, m=0, p=1,R¹¹ is CF₂, and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:

wherein “*” represents the point of attachment to R⁵ (Compound EM).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, n=0, m=0, p=1,R¹¹ is CF₂, and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:

wherein “*” represents the point of attachment to R⁵ (Compound EN).

Another embodiment is a compound having Formula I, wherein R¹═—CH₃,R²═—CH₃, R³═—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, n=0, m=0, p=0,R⁹═—CH₂CH₂CF₃, and R¹⁰═—CH₂CH₂CF₃ (Compound EQ).

Compounds of the present invention include but are not limited to thefollowing:

-   [(3R,4R,5S)-4-(hydroxy)-3-methyl-5-(methylamino)-1-(morpholin-4-yl)hexanoic    acid]¹[(R)-Me-Sar]³cyclosporin A (Compound F);-   [(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-(4-methylpiperazino)-hexanoic    acid]¹[(R)-Me-Sar]³cyclosporin A (Compound L);-   [(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-diethylamino-hexanoic    acid]¹[(R)-Me-Sar]³cyclosporin A (Compound M);-   [(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-(N-3-piperazinone)-hexanoic    acid]¹[(R)-Me-Sar]³cyclosporin A (Compound N);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-{1H-imidazol-4-yl}-ethylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AK);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-methoxyethylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AF);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-methoxyethyl)methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AG);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(N-(3aR*,6aS*)-2-methyloctahydropyrrolo[3,4-c]pyrrolo)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound O);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(1,4-dioxan-2-ylmethyl)methylamino-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AJ);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(thiomorpholino)-hexanoic    acid]¹[(R)-Me-Sar]³cyclosporin A (Compound J);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(1,1-dioxo-thiomorpholino)-hexanoic    acid]¹[(R)-Me-Sar]³cyclosporin A (Compound P);-   [[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-homomorpholino-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound X);-   [(6R,7R,8S)-7-hydroxy-6-methyl-8-(methylamino)-1-N-morpholino-nonanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound G);-   [(6R,7R,8S)-7-Hydroxy-6-methyl-8-(methylamino)-1-N-diethylamino-nonanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AH);-   [(6R,7R,8S)-7-Hydroxy-6-methyl-8-(methylamino)-1-N-(2-methoxy)ethylamino-nonanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AI);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-N-morpholino-1-oxo-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound H);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(4-methylpiperazin-1-yl)-1-oxo-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound T);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-diethylamino-1-oxo-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound U);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-{sulfonic acid    dimethylamide}-ethylamino)-1-oxo-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound W);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-{1H-imidazol-4-yl}-ethylamino)-1-oxo-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound Y);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-({1,1-dioxo}thiomorpholin-4-yl)-1-oxo-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound Z);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(N-(3aR*,6aS*)-2-methyloctahydropyrrolo[3,4-c]pyrrolo)-1-oxo-octanoic    acid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZZ);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-methoxyethylamino)-1-oxo-octanoic    acid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZY);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(1,4-dioxan-2-ylmethyl)amino)-1-oxo-octanoic    acid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZX);-   [(5R,6R,7S)-6-Hydroxy-5-methyl-7-(methylamino)-1-piperidino-1-oxo-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound Q);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-2-(N,N-diethylamino)ethoxy-hexanoic    acid]¹[(S)-thio-isopropyl-Sar]³cyclosporin A (Compound AL);-   [(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoic    acid]¹[(R)-ethyl-Sar]³cyclosporin A (Compound AB);-   [(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoic    acid]¹[(S)-thiomethyl-Sar]³cyclosporin A (Compound K);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoic    acid]¹[(S)-thio-isopropyl-Sar]³cyclosporin A (Compound I);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoic    acid]¹[(S)-methoxy-Sar]³cyclosporin A (Compound B);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoic    acid]¹[(R)-methoxymethylene-Sar]³cyclosporin A (Compound D);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoic    acid]¹[(R)-hydroxymethyl-Sar]³cyclosporin A (Compound V);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoic    acid]¹[(R)-2 diethylamino ethyl oxymethyl-Sar]³cyclosporin A    (Compound S);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-homomorpholino-hexanoic    acid]¹[(R)-2 diethylamino ethyl oxymethyl-Sar]³cyclosporin A    (Compound AD);-   [(5R,6R,7S)-1-(dimethylamino)-6-hydroxy-5-methyl-7-(methylamino)-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound KF);-   [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(N-morpholino)-octanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound KG);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoic    acid]¹[(S)-2-diethylaminoethylthiomethyl-Sar]³cyclosporinA (Compound    AC);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-2-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DA);-   [(3R,4R,5S)-1-(Bis{pyridin-2-ylmethyl}amino)-4-hydroxy-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DB);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(methyl-phenyl-amino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DC);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(methyl-pyridin-2-yl-amino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DD);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-sulfamoyl-ethyl)-methyl-amino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DE);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-3-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DF);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyrimidin-2-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DG);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyrazin-2-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DH);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3-methyl-3H-imidazol-4-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DI);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({2-methyl-2H-pyrazol-3-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DJ);-   [(3R,4R,5S)-1-({2-Cyano-propyl}-methyl-amino)-4-hydroxy-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DK);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-4-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DL);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({1-methyl-1H-pyrazol-4-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DM);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3,3,3-trifluoropropyl}-methyl-amino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DN);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({1-methyl-3-trifluoromethyl-2H-pyrazol-5-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DO);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({5-fluoro-pyridin-2-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DP);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({5-chloro-pyridin-2-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DQ);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3-trifluoromethyl-pyridin-2-ylmethyl}-methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DR);-   [(3R,4R,5S)-1-(3,3-Dimethyl-morpholin-4-yl)-4-hydroxy-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DS);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-methylamino-((R)-3-methyl-morpholin-4-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DT);-   [(3R,4R,5S)-1-(5,6-Dihydro-8H-[1,2,4]triazolo[4,3-a]pyrazin-7-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DU);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(8-oxa-3-aza-bicyclo[3.2.1]oct-3-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DV);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-((S)-3-methyl-morpholin-4-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DW);-   [(3R,4R,5S)-1-(2,3-Dihydro-benzo[1,4]oxazin-4-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DX);-   [(3R,4R,5S)-1-(1,4-Dioxa-8-aza-spiro[4.5]dec-8-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DY);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-phenyl-morpholin-4-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DZ);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(piperidin-1-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EA);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(pyrrolidin-1-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EF);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-trifluoromethyl-piperidin-1-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EB);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(3-trifluoromethyl-morpholin-4-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EC);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-([1,2]oxazinan-2-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound ED);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-trifluoromethyl-5,6-dihydro-8H-imidazo[1,2-a]pyrazin-7-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EE);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(pyrrolidin-1-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EF);-   [[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(4-methyl-[1,4]diazepan-1-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EG);-   [[(3R,4R,5S)-4-Hydroxy-1-(3-methoxy-azetidin-1-yl)-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EH);-   [(4R,5R,6S)-5-Hydroxy-4-methyl-6-(methylamino)-1-(morpholin-4-yl)-heptanoic    acid]¹cyclosporin A (Compound EI);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-1-(morpholin-4-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³[Ethyl-Val]⁴cyclosporin A (Compound EJ);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2,2,6,6-tetrafluoro-morpholin-4-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EK);-   [(3R,4R,5S)-(3,3-Difluoro-pyrrolidin-1-yl)-4-Hydroxy-3-methyl-5-(methylamino)-1-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EL);-   [(3R,4R,5S)-(3,3-Difluoro-azetidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-1-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EM);-   [(3R,4R,5S)-(4,4-Difluoro-piperidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-1-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EN);-   [(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(3,3,4,4-tetrafluoro-pyrrolidin-1-yl)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound ER);-   3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2,2,6,6-tetrafluoro-morpholin-4-yl)-hexanoic    acid]¹[(R)-hydroxymethyl-Sar]³cyclosporin A (Compound EO);-   (3R,4R,5S)-1-(3,3-Difluoro-pyrrolidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-hydroxymethyl-Sar]³cyclosporin A (Compound EP); and-   (3R,4R,5S)-1-[Bis-(3,3,3-trifluoro-propyl)-amino]-[(4-Hydroxy-3-methyl-5-(methylamino)-hexanoic    acid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EQ).

In another embodiment there is provided a pharmaceutical compositioncomprising a compound having Formula I and a pharmaceutically acceptableexcipient. The excipient can be an ophthalmically acceptable excipient.

The pharmaceutical composition can be in the form of a liquid, solid, oremulsion. For example, the pharmaceutical compositions can be in theform of an aqueous solution.

A compound of Formula I can be in a purified form. In one embodiment,the purified form is the form obtained from medium pressure liquidchromatography (MPLC).

The present invention includes pharmaceutically acceptable salts of anycompound having Formula I.

The term “pharmaceutically acceptable salts” refers to salts orcomplexes that retain the desired biological activity of the compound ofFormula I and exhibit minimal or no undesired toxicological effects tothe patient, animal, or cell system to which they are administered. The“pharmaceutically acceptable salts” according to the invention includetherapeutically active non-toxic base or acid salt forms of Formula I.

The acid addition salt form of a compound of Formula I that occurs inits free form as a base can be obtained by treating the free base withan appropriate acid such as an inorganic acid, for example, hydrochloricacid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid andthe like; or an organic acid such as for example, acetic acid,hydroxyacetic acid, propanoic acid, lactic acid, pyruvic acid, malonicacid, fumaric acid, maleic acid, oxalic acid, tartaric acid, succinicacid, malic acid, ascorbic acid, benzoic acid, tannic acid, pamoic acid,citric acid, methylsulfonic acid, ethanesulfonic acid, benzenesulfonicacid, or formic acid and the like (Handbook of Pharmaceutical Salts, P.Heinrich Stahal & Camille G. Wermuth (Eds), Verlag Helvetica ChemicaActa—Zürich, 2002, 329-345).

The base addition salt form of a compound of Formula I that occurs inits free form as an acid can be obtained by treating the acid with anappropriate base such as an inorganic base, for example, sodiumhydroxide, magnesium hydroxide, potassium hydroxide, calcium hydroxide,ammonia and the like; or an organic base such as for example,L-arginine, ethanolamine, betaine, benzathine, morpholine and the like.(Handbook of Pharmaceutical Salts, P. Heinrich Stahal & Camille G.Wermuth (Eds), Verlag Helvetica Chemica Acta—Zürich, 2002, 329-345).

The present invention further concerns the use of a compound of FormulaI, or a pharmaceutically acceptable salt thereof, for the manufacture ofa pharmaceutical composition.

The present invention further encompasses a method for treating amedical condition in a patient in need thereof, the method comprisingadministering to the patient a therapeutically effective amount of acompound of Formula I, or a pharmaceutically acceptable salt thereof. Insome forms of this method, the medical condition is selected from thegroup consisting of dry eye, dry eye disease, ocular surfaceinflammation, blepharitis, meibomian gland disease, allergicconjunctivitis, pterygia, ocular symptoms of graft versus host disease,ocular allergy, atopic keratoconjunctivitis, vernalkeratoconjunctivitis, uveitis, anterior uveitis, Behcet's disease,Stevens Johnson syndrome, ocular cicatricial pemphigoid, chronic ocularsurface inflammation caused by viral infection, herpes simplexkeratitis, adenoviral keratoconjunctivitis, ocular rosacea, orpinguecula.

Another embodiment is a method for reducing corneal transplant rejectionin a patient in need thereof, the method comprising administering to thepatient a therapeutically effective amount of a compound of Formula I,or a pharmaceutically acceptable salt thereof.

Another embodiment is a method for reducing inflammation of the eyecaused by an ocular surgery, the method comprising administering to theeye(s) of a patient who has received ocular surgery a therapeuticallyeffective amount of a compound of Formula I, or a pharmaceuticallyacceptable salt thereof.

Another embodiment is a method for treating dry eye in a patient in needthereof, the method comprising administering to the eye(s) of thepatient a therapeutically effective amount of a compound of Formula I,or a pharmaceutically acceptable salt thereof.

Another embodiment is a method for increasing tear production in apatient whose tear production is suppressed or presumed to be suppresseddue to ocular inflammation associated with keratoconjunctivitis sicca,the method comprising administering to the eye(s) of the patient atherapeutically effective amount of a compound of Formula I, or apharmaceutically acceptable salt thereof.

Another embodiment is a method for reducing or preventing an ocularcondition in a patient, the method comprising administering atherapeutically effective amount of a compound of Formula I to thepatient. In some forms of this method, the ocular condition is selectedfrom the group consisting of dry eye, ocular surface inflammation,blepharitis, meibomian gland disease, allergic conjunctivitis, pterygia,ocular symptoms of graft versus host disease, ocular allergy, atopickeratoconjunctivitis, vernal keratoconjunctivitis, uveitis, anterioruveitis, ocular cicatricial pemphigoid, chronic ocular surfaceinflammation caused by viral infection, herpes simplex keratitis,adenoviral keratoconjunctivitis, ocular rosacea, and pinguecula.

A compound of Formula I may be administered in the form of apharmaceutical composition, topically, orally, systemically, or by othersuitable routes.

Many compounds having Formula I are potent inhibitors of cyclophilin A(CyP-A), and are non-immunosuppressive as measured by the Calcineurinphosphatase assay (IC50>10 μM) and mixed lymphocyte reaction (MLR) assay(>50 fold less active than Cyclosporin A). However, a sub-set ofcompounds of Formula I has been discovered that show immunosuppressiveactivity as measured by the Calcineurin phosphatase assay (IC50<4 μM)and MLR assay (<25 fold less active than Cyclosporin A).

The present invention includes, but is not limited to, the followingembodiments 1-20:

-   1. A compound having Formula I

or a pharmaceutically acceptable salt thereof, wherein:R¹ is —H, —C₁₋₆alkyl, —OC₁₋₆alkyl, —CH₂F, —CH₂OCH₃, —SC₁₋₆alkyl, —CH₃,—CH₂CH₃, —SCH(CH₃)₂, —CH₂OH, —SCH₃, —OCH₃, —R¹³R¹⁴,

R² is —CH₃, —CH₂CH₃, or —CH₂CH₂CH₃;

R³ is —CH₂CH(CH₃)₂, —CH(CH₃)₂, —CH₂C(CH₃)₂(OH), —CH(CH₃)(CH₂CH₃), or—CH₂CH(R⁷)(CH₂CH₃);

R⁴ is —CH₃ or —CH₂OH;

R⁵ is —R⁸(CH₂)_(n)(C═O)_(m)—;

R⁶ is —CH₂CH₃, —CH(CH₃)(OH), —CH(CH₃)₂, or —CH₂CH₂CH₃;

R⁷ is OC₁₋₅ alkyl;

R⁸ is O, S, CH₂O, CH₂S, or CH₂;

R⁹ is —H, —C₁₋₅ alkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or taken togetherwith R¹¹, R¹⁰, and the N to which R⁹ and R¹⁰ are attached forms aheterocycle;R¹⁰ is —H, —C₁₋₅ alkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or taken togetherwith R¹¹, R⁹, and the N to which R⁹ and R¹⁰ are attached forms aheterocycle;R¹¹ is O, NR¹², S(O)_(q), C₁₋₅alkylene, divalent C₃₋₈cycloalkyl,divalent heterocycle, carbonyl, or taken together with R⁹, R¹⁰, and theN to which R⁹ and R¹⁰ are attached forms a heterocycle;R¹² is H, CH₃, or C₁₋₅ alkyl;

R¹³ is O, S, CH₂O, CH₂S, CH₂SO, or CH₂SO₂;

R¹⁴ is H, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, C₃₋₈cycloalkyl,CH₂CH₂N(CH₂CH₃)₂, CH₂CH₂NH(CH₂CH₃), heterocycle, or aryl;n=0, 1, 2, 3, or 4;m=0 or 1;p=0 or 1; andq=0, 1, or 2;wherein R¹⁴ is optionally substituted with one or more groupsindependently selected from the group consisting of H, C₁₋₆alkyl,halogen, hydroxyl, ester, sulfonamide, ketone, aldehyde, cycloalkyl,heterocycle, aryl, amine, heterocycle, amide, and guanidinyl;wherein the heterocycle comprising R⁹, R¹⁰, R¹¹, and the N to which R⁹and R¹⁰ are attached is monocyclic or polycyclic;wherein “- - -” is a single bond or is absent; andwith the provisos thatwhen R⁸ is O, S, CH₂O, or CH₂S then n is not 0 or 1;when p=0 then R¹¹ and “- - -” are absent; andwhen R¹¹ and “- - -” are absent then R⁹ is not directly linked to R¹⁰.

-   2. A compound according to embodiment 1, above, wherein R¹ is not    hydrogen (H).-   3. A compound according to embodiment 1, wherein when R¹ is H then    m=0.-   4. A compound according to embodiment 1, wherein when R¹ is H, then    m=0 and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached    together form a heterocycle.-   5. A compound according to embodiment 1, wherein R² is —CH₃ or    —CH₂CH₃; R³ is —CH₂CH(CH₃)₂, —CH(CH₃)₂, or —CH₂C(CH₃)₂(OH); R⁴ is    —CH₃; R⁶ is —CH₂CH₃; and R⁸ is CH₂.-   6. A compound according to embodiment 1, wherein R² is —CH₃; R³ is    —CH₂CH(CH₃)₂; R⁴ is —CH₃; R⁶ is —CH₂CH₃; and R⁸ is CH₂.-   7. A compound according to embodiment 1, wherein R¹ is —CH₃,    —CH₂CH₃, or —SCH₃; R² is —CH₃; R³ is —CH₂CH(CH₃)₂; R⁴ is —CH₃; R⁶ is    —CH₂CH₃; and R⁸ is CH₂.-   8. A compound according to embodiment 1, wherein the compound is any    one of those listed in Tables 1-20.-   9. A pharmaceutical composition comprising as active ingredient a    therapeutically effective amount of a compound according to any one    of embodiments 1-8, or a pharmaceutically acceptable salt thereof,    and a pharmaceutically acceptable excipient.-   10. The pharmaceutical composition of embodiment 9, wherein the    compound is present in the composition in an amount of from about    0.01% (w/v) to about 1% (w/v).-   11. The pharmaceutical composition of embodiment 10, wherein the    composition is in the form of an aqueous solution.-   12. The pharmaceutical composition of embodiment 11, wherein the    composition is acceptable for administration to the eye(s) of a    mammal.-   13. A method of treating a medical condition in a patient in need    thereof, the method comprising administering to the patient a    therapeutically effective amount of a compound, or a    pharmaceutically acceptable salt thereof, according to any one of    embodiments 1-8.-   14. The method of embodiment 13, wherein the patient is a human    patient.-   15. The method of embodiment 13, wherein the medical condition is    dry eye, dry eye disease, ocular surface inflammation, blepharitis,    meibomian gland disease, allergic conjunctivitis, pterygia, ocular    symptoms of graft versus host disease, ocular allergy, atopic    keratoconjunctivitis, vernal keratoconjunctivitis, uveitis, anterior    uveitis, Behcet's disease, Steven Johnson syndrome, ocular    cicatricial pemphigoid, chronic ocular surface inflammation caused    by viral infection, herpes simplex keratitis, adenoviral    keratoconjunctivitis, ocular rosacea, or pinguecula.-   16. A method for reducing corneal transplant rejection in a patient    in need thereof, the method comprising administering to the patient    a therapeutically effective amount of a compound, or a    pharmaceutically acceptable salt thereof, according to any one of    embodiments 1-8.-   17. A method for reducing inflammation of the eye caused by an    ocular surgery, the method comprising administering to the eye(s) of    a patient who has received ocular surgery a therapeutically    effective amount of a compound, or a pharmaceutically acceptable    salt thereof, according to any one of embodiments 1-8.-   18. A method for treating dry eye in a patient in need thereof, the    method comprising administering to the eye(s) of the patient a    therapeutically effective amount of a compound, or a    pharmaceutically acceptable salt thereof, according to any one of    embodiments 1-8.-   19. A method for increasing tear production in a patient whose tear    production is suppressed or presumed to be suppressed due to ocular    inflammation associated with keratoconjunctivitis sicca, the method    comprising administering to the eye(s) of the patient a    therapeutically effective amount of a compound according to any one    of embodiments 1-8, or a pharmaceutically acceptable salt thereof.-   20. The method of any one of embodiment 13, wherein the compound is    administered to the patient topically, orally, or systemically.    The present invention further includes a method for making a    compound having the formula:

the method comprising

-   -   a) adding 10% palladium on carbon to a solution comprising

in ethanol, wherein P is a protecting group;

-   -   b) stirring the solution under a hydrogen atmosphere;    -   c) filtering the mixture from step b through a filter and        collecting the filtrate;    -   d) washing the filter used in step c and collecting the filtrate        together with the filtrate from step c;    -   e) evaporating the filtrate from step d to thereby obtain a        compound having the formula shown above as the major product.        In some forms of this method for making, P is

and the filter comprises celite. In yet more specific forms of thismethod washing the filter used in step c comprises washing the filterwith ethyle acetate.

DEFINITIONS

A “patient in need in need of treatment” or “patient in need thereof”refers to a human or non-human mammal afflicted with a medicalcondition, as specified in context. Non-limiting examples of a non-humanmammal include a horse, pig, monkey, dog, rabbit, guinea pig, rat, ormouse.

A “therapeutically effective amount” refers to the amount of a compoundsufficient to reduce the severity of one or more symptoms associatedwith, accompanying, or resulting from a medical condition affecting asubject.

“Treating” and “to treat” refers to relieving or reducing at least onesymptom associated with or accompanying a medical condition. Forexample, treatment of dry eye and relief of inflammation of the ocularsurface, as may occur in an individual suffering from dry eye, may beobserved or experienced as an improvement in vision, and/or as areduction in swelling, pain, redness, dryness, scratchiness, grittiness,foreign body sensation, stinging, burning, or itching. Treating aninflammation of the ocular surface or ocular surface adnexa may improvethe visual performance and the optical quality of the eye. Improvementin visual performance may include improved optical quality, improvedtear film production, secretion, quality, and/or stability, reducedblurring, improved central and/or peripheral field vision, improvedvisual performance, acuity, or perception, and/or reduced blinkingfrequency. The symptom(s) positively affected by the treatment, willdepend on the particular condition.

The term “inflammation” refers to the biological response of the livingbody to injury or other harmful insults. Symptoms of “an inflammation atthe ocular surface” can include redness, swelling, heat, pain, and/orloss of function of glands or tissue in the ocular surface or ocularsurface adnexa. Other symptoms may include sensations of (and lead apatient to complain of) dryness, burning, itching, or scratchiness. Asubject may report a feeling of dust, dirt, sand, or gravel in the eye.

A “Medical condition” refers to a deviation from or interruption of thenormal structure or function of any body part, tissue, organ, or systemand that is characterized by an identifiable group of signs or symptomswhose etiology, pathology, and prognosis may be known or unknown. Amedical condition of a body part, tissue, organ, or system of a human ornon-human mammal may result from various causes, including but notlimited to injury, surgical trauma, infection, nutritional deficiency,genetic defect, exposure to toxins or radiation, and environmentalstress. Medical conditions include ocular conditions such as, forexample, inflammation of the ocular surface, and dry eye; anddermatological conditions such as an inflammation of the skin.

An “ocular condition” is a disease, ailment or condition which affectsor involves the eye or one or more parts or regions of the eye.

“Ocular surface condition” refers to a medical condition that affects orinvolves one or more parts, regions, or tissues of the ocular surface.An ocular surface condition can be an inflammation of an ocular surfacetissue, and includes an acute, chronic, and surgically-inducedinflammation of an ocular surface tissue.

The term “ocular surface” refers to the cornea, the corneal epithelium,the conjunctiva (palpebral, bulbar, and formiceal), the conjunctivalblood vessels, Tenon's capsule, the sclera, and the limbus.

The term “ocular surface adnexa” refers to structures in close proximityto the ocular surface, including the lacrimal gland, the eye lids,eyelashes, and eyebrows, the orbital wall, the periocular or extraocularmuscles, and the meibomian glands.

The “eye” is the sense organ for sight, and includes the eyeball, orglobe, the orbital sense organ that receives light and transmits visualinformation to the central nervous system. Broadly speaking the eyeincludes the eyeball and the tissues and fluids which constitute theeyeball, the periocular muscles (such as the oblique and rectus muscles)and the portion of the optic nerve which is within or adjacent to theeyeball.

The “eye lids” are the structures covering the front of the eye thatprotect it, limit the amount of light entering the pupil, and helpdistribute tear film over the exposed corneal surface.

The term “biocompatible” means compatible with living tissue or a livingsystem by not being toxic, injurious, or physiologically reactive and bycausing minimal or no immunological reaction.

The term “alkyl”, as used herein, refers to saturated monovalent ordivalent hydrocarbon moieties having linear or branched moieties orcombinations thereof and containing 1 to 6 carbon atoms. One methylene(—CH₂—) group, of the alkyl can be replaced by oxygen, sulfur,sulfoxide, nitrogen, carbonyl, carboxyl, sulfonyl, amide, sulfonamide,by a divalent C₃₋₆ cycloalkyl, by a divalent heterocycle, or by adivalent aryl group. Alkyl groups can be independently substituted byhalogen, hydroxyl, cycloalkyl, amine groups, heterocyclic groups,carboxylic acid groups, phosphonic acid groups, sulphonic acid groups,phosphoric acid groups, nitro groups, amide groups, or sulfonamidesgroups. Non-limiting examples of suitable alkyl groups include methyl(—CH₃), ethyl (—CH₂CH₃), n-propyl (—CH₂CH₂CH₃), isopropyl (—CH(CH₃)₂),and t-butyl (—C(CH₃)₃).

An “alkylene” is a divalent alkyl. Non-limiting examples of an alkyleneinclude methylene, ethylene (—CH₂CH₂—), and n-propylene (—CH₂CH₂CH₂—).

The term “cycloalkyl”, as used herein, refers to a monovalent ordivalent group of 3 to 8 carbon atoms derived from a saturated cyclichydrocarbon. Cycloalkyl groups can be monocyclic or polycyclic.Cycloalkyl can be independently substituted by halogen, nitro groups,cyano groups, —OC₁₋₆ alkyl groups, —SC₁₋₆ alkyl groups, —C₁₋₆ alkylgroups, —C₂₋₆ alkenyl groups, —C₂₋₆ alkynyl groups, C₃₋₈ cycloalkylgroups, carboxylic acid groups, ester groups, ketone groups, aldehydegroups, amide groups, amine groups, sulfonamide groups or hydroxylgroups.

The term “cycloalkenyl”, as used herein, refers to a monovalent ordivalent group of 3 to 8 carbon atoms derived from a saturatedcycloalkyl having at least one double bond. Cycloalkenyl groups can bemonocyclic or polycyclic. Cycloalkenyl groups can be independentlysubstituted by halogennitro groups, cyano groups, —OC₁₋₆ alkyl groups,—SC₁₋₆ alkyl groups, —C₁₋₆ alkyl groups, —C₂₋₆ alkenyl groups, —C₂₋₆alkynyl groups, carboxylic acid groups, ester groups, ketone groups,aldehyde groups, amide groups, amine groups, sulfonamide groups, C₃₋₈cycloalkyl groups or hydroxyl groups.

The term “halogen”, as used herein, refers to an atom of chlorine,bromine, fluorine, iodine.

The term “haloalkyl” refers to an alkyl in which one or more hydrogenatoms on the alkyl have been replaced with a halogen atom. Non-limitingexamples of a haloalkyl include fluoroalkyls such as —CF₃ and—CH₂CH₂CF₃.

The term “heterocycle” as used herein, refers to a 3 to 10 memberedring, which can be aromatic or non-aromatic, monvalent or divalent,saturated or unsaturated, containing at least one heteroatom selectedform O or N or S or combinations of at least two thereof, interruptingthe carbocyclic ring structure. The heterocyclic ring can be interruptedby a C═O; the S and N heteroatoms can be oxidized. Heterocycles can bemonocyclic or polycyclic. For example, a heterocyle can be bicyclic. Therings in a bicyclic or polycyclic heterocycle can be fused or non-fused.Heterocyclic ring moieties can be substituted by halogen, nitro groups,cyano groups, —OC₁₋₆ alkyl groups, —SC₁₋₆ alkyl groups, —C₁₋₆ alkylgroups, —C₂₋₆ alkenyl groups, —C₂₋₆ alkynyl groups, carboxylic acidgroups, ester groups, ketone groups, aldehyde groups, amide groups,amine groups, sulfonamide groups, C₃₋₈ cycloalkyl groups, or hydroxylgroups.

The term “alkenyl”, as used herein, refers to a monovalent or divalenthydrocarbon radical having 2 to 6 carbon atoms, derived from a saturatedalkyl, having at least one double bond. A C₂₋₆ alkenyl can be in the Eor Z configuration. Alkenyl groups can be substituted by C₁₋₃ alkyl, asdefined above, or by halogen.

The term “alkynyl”, as used herein, refers to a monovalent or divalenthydrocarbon radical having 2 to 6 carbon atoms, derived from a saturatedalkyl, having at least one triple bond. Alkynyl groups can besubstituted by C₁₋₃ alkyl, as defined above, or by halogen.

The term “aryl” as used herein, refers to an organic moiety derived froman aromatic hydrocarbon consisting of a ring containing 6 to 10 carbonatoms by removal of one hydrogen, which can be substituted by halogen,nitro groups, cyano groups, —OC₁₋₆ alkyl groups, —SC₁₋₆ alkyl groups,—C₁₋₆ alkyl groups, —C₂₋₆ alkenyl groups, —C₂₋₆ alkynyl groups,carboxylic acid groups, ester groups, ketone groups, aldehyde groups,amide groups, amine groups, sulfonamide groups, C₃₋₈ cycloalkyl groupsor hydroxyl groups. A non-limiting example of an aryl is phenyl.Preferred substitution site on aryl are the meta and the para positions.Most preferred substitution sites on aryl are the para positions.

The term “ketone” as used herein, represents an organic compound havinga carbonyl group linked to a carbon atom such as —(CO)R^(x) whereinR^(x) can be alkyl, aryl, cycloalkyl, cycloalkenyl, heterocycle asdefined herein.

The term “aldehyde” as used herein, represents a group of formula—C(O)H.

The term “ester” as used herein, represents a group of formula—C(O)OR^(x), wherein R^(x) can be alkyl, aryl, cycloalkyl, cycloalkenyl,heterocycle as defined herein.

The term “hydroxyl” as used herein, represents a group of formula —OH.

The term “carbonyl” as used herein, represents a group of formula—C(O)—, which may also be represented as and is equivalent to —(C═O)—.

The term “carboxyl” as used herein, represents a group of formula—C(O)O—.

The term “sulfonyl” as used herein, represents a group of formula —SO₂—.

The term “sulfate” as used herein, represents a group of formula—O—S(O)₂—O—.

The term “carboxylic acid” as used herein, represents a group of formula—C(O)OH.

The term “nitro” as used herein, represents a group of formula —NO₂.

The term “cyano” as used herein, represents a group of formula —CN.

The term “phosphonic acid” as used herein, represents a group of formula—P(O)(OH)₂.

The term “phosphoric acid” as used herein, represents a group of formula—OP(O)(OH)₂.

The term “amide” as used herein, represents a group of formula—C(O)NR^(x)R^(y), wherein R^(x) and R^(y) can be the same orindependently H, alkyl, aryl, cycloalkyl, cycloalkenyl, or heterocycle,as defined above.

The term “amine” as used herein, represents a group of formula—NR^(x)R^(y), wherein R^(x) and R^(y) can be the same or independentlyH, alkyl, aryl, cycloalkyl, cycloalkenyl, or heterocycle as definedabove.

The term “sulfonamide” as used herein, represents a group of formula—S(O)₂NR^(x)R^(y) wherein R^(x) and R^(y) can be the same orindependently H, alkyl, aryl, cycloalkyl, cycloalkenyl, heterocycle asdefined above.

The term “sulfoxide” as used herein, represents a group of formula—S(O)—.

The term “sulphonic acid” as used herein, represents a group of formula—S(O)₂OH.

The formula “H”, as used herein, represents a hydrogen atom.

The formula “O”, as used herein, represents an oxygen atom.

The formula “N”, as used herein, represents a nitrogen atom.

The formula “S”, as used herein, represents a sulfur atom.

The term “piperazinonyl” as used herein represents a group of formula

The term “morpholinyl” as used herein represents a group of formula

The term “N-methylpiperazinyl” as used herein represents a group offormula

The term “guanidinyl” as used herein refers to a group of formula “

The term “substituted” means that one or more hydrogens on thedesignated atom is replaced with a selection from the indicated group,provided that the designated atom's normal valency under the existingcircumstances is not exceeded, and that the substitution results in astable compound.

The term “optionally substituted” means optional substitution with thespecified groups, radicals or moieties.

The term “purified”, “in purified form” or “in isolated and purifiedform” for a compound refers to the physical state of said compound afterbeing isolated from a synthetic process (e.g. from a reaction mixture),or natural source or combination thereof. Thus, the term “purified”, “inpurified form” or “in isolated and purified form” for a compound refersto the physical state of said compound after being obtained from apurification process or processes described herein or well known to theskilled artisan (e.g., chromatography, recrystallization and the like)in sufficient purity to be characterizable by standard analyticaltechniques described herein or well known to the skilled artisan.

It should also be noted that any carbon as well as heteroatom withunsatisfied valences in the text, schemes, examples and Tables herein isassumed to have the sufficient number of hydrogen atom(s) to satisfy thevalences. And any one or more of these hydrogen atoms can be deuterium.

When a functional group in a compound is termed “protected”, this meansthat the group is in modified form to preclude undesired side reactionsat the protected site when the compound is subjected to a reaction.Suitable protecting groups will be recognized by those with ordinaryskill in the art and in standard textbooks such as, for example, T. W.Greene et al, Protective Groups in organic Synthesis (1981), Wiley, NewYork.

DETAILED DESCRIPTION OF THE INVENTION

Therapeutic Uses

The present invention is directed in part to pharmaceutical compositionscomprising a compound having Formula I.

A pharmaceutical composition comprising a compound of Formula I may beuseful for treating dry eye, dry eye disease (i.e., keratoconjunctivitissicca), ocular surface inflammation (i.e, inflammation of the ocularsurface), blepharitis, meibomian gland disease, allergic conjunctivitis,pterygia, ocular symptoms of graft versus host disease, ocular allergy,atopic keratoconjunctivitis, vernal keratoconjunctivitis, uveitis,anterior uveitis, Behcet's disease, Stevens Johnson syndrome, ocularcicatricial pemphigoid, chronic ocular surface inflammation caused byviral infection, herpes simplex keratitis, adenoviralkeratoconjunctivitis, ocular rosacea, and pinguecula, and to prevent orreduce the risk of corneal transplant rejection in a patient or subjectin need thereof. Additionally, one embodiment of this invention is amethod for administering a pharmaceutical composition of this inventionto a patient before, during, or after ocular surgery (such as refractivesurgery) to reduce and/or prevent inflammation of the eye or ocularsurface caused by the surgery.

Pharmaceutical compositions of the invention may be useful for treatingan inflammation of the ocular anterior segment of the eye. Morespecifically, the pharmaceutical compositions of the invention may beuseful for treating an inflammation of the ocular surface or ocularsurface adnexa.

In addition a pharmaceutical composition comprising a compound ofFormula I may be useful for reducing one or symptoms associated with aninflammatory dermatological condition. One example of an inflammatorydermatological condition that may be subject to treatment is psoriasis.Additionally, a pharmaceutical composition comprising a compound ofFormula I may be useful for treating a viral infection. Examples ofviral infections may include Hepatitis C infection and Hepatitis Binfection.

Accordingly, one embodiment is a method for reducing one or more orsymptoms associated with an inflammatory dermatological condition in apatient in need thereof, comprising administering a pharmaceuticalcomposition comprising a compound of Formula I to the patient. Anotherembodiment is a method for reducing or preventing an inflammatorydermatological condition in a patient, comprising administering atherapeutically effective amount of a compound of Formula I to thepatient. The method may reduce one or more signs or symptoms of theinflammatory dermatological condition. One example of an inflammatorydermatological condition is psoriasis.

Another embodiment is a method for treating an inflammation of the skinin a patient in need thereof comprising administering a pharmaceuticalcomposition comprising a compound of Formula I to the patient. Anotherembodiment is a method for reducing an inflammation of the skin in apatient, comprising administering a therapeutically effective amount ofa compound of Formula I to the patient.

Another embodiment is a method for treating a viral infection in apatient in need thereof, comprising administering a pharmaceuticalcomposition comprising a compound of Formula I to the patient. Anotherembodiment is a method for reducing one or more signs or symptoms of aviral infection, or for inhibiting the progress of a viral infection ina patient, comprising administering a pharmaceutical compositioncomprising a compound of Formula I to the patient.

One embodiment is a method for treating dry eye in a patient in needthereof, comprising administering a pharmaceutical compositioncomprising a compound of Formula I to the eye(s) of the patient.

Another embodiment is a method for increasing tear production in apatient whose tear production is suppressed (or presumed to besuppressed) due to ocular inflammation associated withkeratoconjunctivitis sicca.

Another embodiment is a method for reducing ocular surface inflammationin a patient, comprising administering a therapeutically effectiveamount of a compound having Formula I to the patient. In one method, theocular surface inflammation is associated with keratoconjunctivitissicca. In some forms of this method, the compound having Formula I isadministered topically to the patient's eye(s).

One embodiment is a method for reducing or preventing an ocularcondition in a patient, the method comprising administering atherapeutically effective amount of a compound of Formula I to thepatient. The method may reduce one or more signs or symptoms of theocular condition.

“Dry eye” as used herein includes “dry eye disease” as defined by theInternational Dry Eye Workshop (DEWS) in Lemp et al. (2007) “TheDefinition and Classification of Dry Eye Disease: Report of theDefinition and Classification Subcommittee of the International Dry EyeWorkshop” Ocul. Surf 5:75-92. The International Dry Eye Workshop (DEWS)defines dry eye disease as “a multifactorial disease of the tears andocular surface that results in symptoms of discomfort, visualdisturbance, and tear film instability with potential damage to theocular surface, accompanied by increased osmolarity of the tear film andinflammation of the ocular surface.”

The term “dry eye disease” is considered to be synonymous with “dry eyesyndrome” and “keratoconjunctivitis sicca.” Dry eye disease includes theaqueous deficient (Sjogren and non-Sjogren) and evaporative categoriesof dry eye disease. An individual with dry eye disease may present withsymptoms of both aqueous deficiency (e.g., insufficient tear production)and excessive evaporation of the tear film.

The pharmaceutical composition can be administered to a patienttopically, orally, or systemically (including intravenously orintraarterially). Administration may be to the eye, such as the surfaceof the eye.

The actual amount of the compound to be administered in any given casewill be determined by a physician taking into account the relevantcircumstances, such as the severity of the condition, the age and weightof the patient, the patient's general physical condition, the cause ofthe condition, and the route of administration.

Accordingly, the present invention includes methods for treating any ofthe above ocular conditions in a patient in need thereof byadministering a pharmaceutical composition comprising a compound ofFormula I to the patient. The composition can be administered directlyto the ocular surface of the eye or to an ocular region in the eye.Modes of direct administration to the eye can include topical deliveryand intraocular injection.

Pharmaceutical compositions of the invention may also be useful forrestoring corneal sensitivity that has been impaired due to surgery onthe cornea or other surface of the eye. Impaired corneal sensitivity mayalso occur after viral infection, such as by HSV-1, HSV-2, and VZVviruses. Patients with impaired corneal sensitivity often complain thattheir eyes feel dry, even though tear production and evaporation may benormal, suggesting that “dryness” in such patients may actually be aform of corneal neuropathy that results when corneal nerves are severedby surgery or inflamed after viral infection.

A patient in need of treatment of an “ocular surface inflammation” ormore specifically “dry eye” may complain of superficial scratchy pain,abrasiveness, eye dryness, foreign body sensation, scratchiness, oculardiscomfort, ocular pain, burning, itching, decreased vision, visualblurriness or cloudiness, irritation or pain from bright light, ordecreased visual acuity. Dryness may be experienced and reported as afeeling that moisture is absent, foreign body sensation, and/or as afeeling of dust, sand, or gravel in the eye. Accordingly, a patient withdry eye may experience one or more of the following symptoms: stingingand/or burning, dryness, sensation of foreign body (gritty or sandyfeeling), itching, sensitivity to light, pain or soreness, intermittentblurred vision, tired or fatigued eyes, and frequent blinking

Dry eye may be due to inadequate tear production, a disruption in tearsecretion, decreased tear film quality, or excessive evaporation of thetear film at the ocular surface, any and all of which can lead tosensations of dry eye and eye dryness and/or be associated with dry eyedisease.

An individual having dry eye may exhibit one or more of thecharacteristics or symptoms associated with dry eye disease(keratoconjunctivitis sicca). Methods for diagnosing and monitoring dryeye disease may include those described in Bron et al. (2007)“Methodologies to Diagnose and Monitor Dry Eye Disease:Report of theDiagnostic Methodology Subcommittee of the International Dry EyeWorkShop (2007)” Ocul. Surf. 5(2):108-152, and can include, but are notnecessarily limited to, symptom questionnaires developed for use in dryeye diagnosis, the fluorescein tear film break up test, ocular surfacestaining grading with fluorescein/yellow filter, the Schirmer test, andtear osmolarity measurement.

A common feature of dry eye disease is an unstable tear film due toabnormal or deficient tear production, increased tear evaporation, orimbalance of tear components. An unstable tear film may lead to orpromote inflammation of the ocular surface (Pflugfelder et al. 2004, Am.J. Ophthalmol. 137:337-342).

An individual suffering from or in need of treatment of “dry eye,” forthe present invention, can be one that presents with, is suffering from,or exhibits one or more symptoms of dry eye disease, or ocular surfacedryness, or eye dryness, which depending on the individual may includesensations of dry eye (i.e., sensations of eye dryness), tear filminstability, decreased tear secretion, delayed clearance, and alteredtear composition, or tear hyperosmolarity.

For purposes of the present invention, “dry eye” that may potentially betreated with the present pharmaceutical compositions may be chronic ortemporary, may occur in one or both eyes of an individual, and inparticular patients may be due to or caused by changes in physiologicalcondition; use of contact lenses; allergy to a medication; in responseto an external environmental factor such as pollen, dust, particulates,or low humidity; due to a side effect of a medication; aging; low blinkrate; vitamin A deficiency; a chemical burn; radiation; blepharitis;rosacea; reaction to the use preservative-containing topical eye drops,such as wetting drops; disorders of the lid aperature; meibomian oildeficiency; lacrimal deficiency; disruption or damage of the lacrimalgland or obstruction of the lacrimal gland duct; reflex block;infection; changes in hormonal balance; eye surgery, including but notlimited to refractive laser eye surgery, including LASIK, LASEK, andPRK; or as a result of exposure to an environmental contaminantencountered during a recreational or occupational activity; or as aresult of physical injury to the eye. Accordingly, pharmaceuticalcompositions of the present invention may reduce the severity of one ormore symptoms associated with or accompanying dry eye.

In particular patients suffering from dry eye disease, to which thepresent method may be directed, the dry eye disease may be caused bynutritional disorders or deficiencies (including vitamins),pharmacologic side effects, eye stress and glandular and tissuedestruction, environmental exposure to smog, smoke, excessively dry air,airborne particulates, autoimmune and immunodeficient disorders, and maybe prevalent in patients who are unable to blink.

In other forms, the present invention may be directed to treating dryeye associated with rheumatoid arthritis, lupus erythematosus,polymyositis, rosacea, scleroderma, polyarteritis, thyroiditis,hepatobiliary disease, lymphoma, pulmonary fibrosis, macroglobulinemia,or coeliac disease.

Blepharitis is a disorder of the meibomian glands, which produce thelipid layer of tear film. With blepharitis, the glands may becomeinflamed. Symptoms of blepharitis may include eye irritation, soreness,redness and an accumulation of matter on the eyelids. Patients may alsoexperience dry eye as well. Patients suffering from blepharitis maycomplain of a sandy or itchy feeling of their eyes. There is usuallyredness, thickening, and irregularity of the lid margins. Accordingly,blepharitis involves an inflammation of the eye lid margins. Blepharitiscan also affect the conjunctiva, tear film, and the corneal surface inadvanced stages and may be associated with dry eye. Blepharitis iscommonly classified into anterior or posterior blepharitis, withanterior affecting the lash bearing region of the lids, and posteriorprimarily affecting the meibomian gland orifices.

Meibomian gland disease most often occurs as one of three forms: primarymeibomitis, secondary meibomitis, and meibomian seborrhea. Meibomianseborrhea is characterized by excessive meibomian secretion in theabsence of inflammation (hypersecretory meibomian gland disease).Primary meibomitis, by contrast, is distinguished by stagnant andinspissated meibomian secretions (obstructive hypersecretory meibomiangland disease). Secondary meibomitis represents a localized inflammatoryresponse in which the meibomian glands are secondarily inflamed in aspotty fashion from an anterior lid margin blepharitis.

Impaired corneal sensitivity often occurs after refractive surgery, suchas photorefractive keratectomy, laser assisted sub-epitheliumkeratomileusis (LASEK), EPI-LASEK, customized transepithelialnon-contact ablation, or other procedures in which the corneal nervesare severed. Impaired corneal sensitivity may also occur after viralinfection, such as by HSV-1, HSV-2, and VZV viruses. Patients withimpaired corneal sensitivity often complain that their eyes feel dry,even though tear production and evaporation may be normal, suggestingthat “dryness” in such patients may actually be a form of cornealneuropathy that results when corneal nerves are severed by surgery orinflamed after viral infection.

Allergic conjunctivitis is an inflammation of the conjunctiva resultingfrom hypersensitivity to one or more allergens. It may be acute,intermittent, or chronic. It occurs seasonally, that is, at only certaintime of the year, or it occurs perennially, that is, chronicallythroughout the year. Symptoms of seasonal and perennial allergicconjunctivitis include, in addition to inflammation of the conjunctiva,lacrimation, tearing, conjunctival vascular dilation, itching, papillaryhyperlasia, chemosis, eyelid edema, and discharge from the eye. Thedischarge may form a crust over the eyes after a night's sleep.

Atopic keratoconjunctivitis is a chronic, severe form of allergicconjunctivitis that often leads to visual impairment. Symptoms includeitching, burning, pain, redness, foreign body sensation, lightsensitivity and blurry vision. There is often a discharge, especially onawakening from a night's sleep; the discharge may be stringy, ropy, andmucoid. The lower conjunctiva is often more prominently affected thanthe upper conjunctiva. The conjunctiva may range from pale, edematous,and featureless to having the characteristics of advanced disease,including papillary hypertrophy, subepithelial fibrosis, formixforeshortening, trichiasis, entropion, and madurosis. In some patientsthe disease progresses to punctate epithelial erosions, cornealneovascularization, and other features of keratopathy which may impairvision. There is typically goblet cell proliferation in the conjunctiva,epithelial pseudotubular formation, and an increased number ofdegranulating eosinophils and mast cells in the epithelium. CD25+Tlymphocytes, macrophages, and dendritic cells (HLA-DR.sup.+, HLA-CD1+)are significantly elevated in the substantia propria.

Like atopic keratoconjunctivitis, vernal keratoconjunctivitis is asevere form of allergic conjunctivitis, but it tends to affect the upperconjunctiva more prominently than the lower. It occurs in two forms. Inthe palpebral form, square, hard, flattened, closely packed papillae arepresent; in the bulbar (limbal) form, the circumcorneal conjunctivabecomes hypertrophied and grayish. Both forms are often accompanied by amucoid discharge. Corneal epithelium loss may occur, accompanied by painand photophobia, as may central corneal plaques and Trantas' dots.

Uveitis, the inflammation of the uvea, is responsible for about 10% ofthe visual impairment in the United States. Phacoanaphylacticendophthalmitis is a human autoimmune disease. Panuveitis refers toinflammation of the entire uveal (vascular) layer of the eye. Posterioruveitis generally refers to chorioentinitis, and anterior uveitis refersto iridocyclitis.

The inflammatory products (i.e. cells, fibrins, excess proteins) ofthese inflammations are commonly found in the fluid spaces if the eye,i.e. anterior chamber, posterior chamber and vitreous space as well asinfiltrating the tissue intimately involved in the inflammatoryresponse. Uveitis may occur following surgical or traumatic injury tothe eye; as a component of an autoimmune disorder, such as rheumatoidarthritis, Behcet's disease, ankylosing spondylitis, and sarcoidosis; asan isolated immune mediated ocular disorder, such as pars planitis,iridocyclitis etc., unassociated with known etiologies; and followingcertain systemic diseases which cause antibody-antigen complexes to bedeposited in the uveal tissues. Together these disorders represent thenon-infectious uveitities.

Phacoanaphylaxis is a severe form of uveitis in which the lens in thecausative antigen. The lens proteins are normally secluded by the lenscapsule since before birth. When these proteins are released into theeye by injury or by surgery or occasionally during cataract development,they can become intensely antigenic and incite an autoimmune response.If the response is moderate it is seen as chronic uveitis. If it is veryfast in progression the eye becomes seriously inflamed in all segments.This latter response is named phacoanaphylaxis.

Uveitis is a prominent feature of Behcet's disease, a multi-systeminflammatory disorder also characterized by oral and genital ulcers,cutaneous, vascular, joint, and neurological manifestations

Rosacea is a chronic and common skin disorder with no identified causeor cure. The pathogenesis of rosacea is thought to have multiplefactors. Possible factors include exposure to the demodex folliculorummite, gastrointestinal disease or a vasodilation disorder, and othertriggers such as diet or sunlight. Patients may present with a varietyof symptoms, including inflammatory papules, edema, telangiectasia,rhinophyma and ocular symptoms.

The ocular signs of rosacea include blepharitis, including anteriorblepharitis, conjunctivitis, iritis, iridocyclitis, keratitis, meibomiangland dysfunction, telangiectasia, erythema, chalazion, hordeolum,interpalpebral hyperemia, conjuctival hyperemia, ciliary base injection,bulbar injection, crusts, sleeves, and superficial punctuatekeratopathy. The ocular symptoms are nonspecific and may includeburning, tearing, decreased tear secretion, redness, and foreign body orgritty or dry sensation, irritation, Itchiness, Blurred vision,Photosensitivity, Watery eyes, bloodshot eyes, Burning, telangiectasia,irregularity of the lid margins, and meibomian gland dysfunction.

Pinguecula is a benign, yellowish brown proliferative growth that formson the conjunctiva. Pinguecula may cause irritation and scratchiness ofthe eye, dry eye, inflammation of the conjunctiva and effect appearanceof the eye. Inflamed pinguecula, which cause ocular irritation or becomeunsightly, may require surgical removal. However, the post-operationscar may be as cosmetically objectionable as the pinguecula andpinguecula regrowth may occur following surgical removal.

Allogeneic bone marrow transplantation (BMT) is a well-establishedtreatment for malignant and non-malignant hematological diseases, and isperformed in tens of thousands of patients each year. Mature donor Tcells within the stem cell graft are the main mediators of thebeneficial immune effects, but they are also responsible for theinduction of graft-versus-host disease (GVHD), the major cause ofmorbidity and mortality in BMT patients. GVHD occurs when transplanteddonor-derived T cells recognize proteins expressed by recipientantigen-presenting cells. Consequently, this recognition induces donorT-cell activation, proliferation, and differentiation, leading to acellular and inflammatory attack on recipient target tissues. Acute orchronic GVHD occurs within a 100-day period post-BMT that leads todermatitis, enteritis, and hepatitis. Ocular symptoms include blurryvision, foreign body sensation, burning sensation, severe lightsensitivity, chronic conjunctivitis, dry eye, and eye pain.

Pharmaceutical Compositions

The present invention includes pharmaceutical compositions comprising,consisting of, or consisting essentially of a compound having Formula I,or a pharmaceutically acceptable salt thereof, in combination with oneor more pharmaceutically acceptable excipients. A pharmaceuticallyacceptable excipient may improve the stability or effectiveness of thecomposition. A “pharmaceutically acceptable excipient” is one that iscompatible with the compound of Formula I and that is not harmful to theperson receiving the pharmaceutical composition. Mixtures of two or moreof such suitable excipients may be used. A pharmaceutical compositionmay comprise two or more compounds having Formula I, or two or moresalts thereof.

Pharmaceutical compositions of the present invention can be in the formof a liquid (such as an aqueous solution), solid, gel, or emulsion. Thecomposition can be sterilized and therefore prepared in sterile form forpharmaceutical use.

The pharmaceutical composition may be prepared in a unit dosage formsuitable for oral, systemic (arterial or intravenous), or topicaladministration to a patient. For example the pharmaceutical compositionmay be prepared in an aqueous liquid or emulsion form suitable oracceptable for administration or topical application to the eye(s) ofthe patient.

For topical ocular applications (such as administration to the eye),pharmaceutical compositions may be prepared by combining atherapeutically effective amount of a compound of Formula I, or apharmaceutically acceptable salt thereof, as an active ingredient, withone or more pharmaceutically acceptable excipients. For ocularapplications, the excipient is further preferably ophthalmicallyacceptable, that is, is causes little or no injury to the eye.

A therapeutically effective amount of a compound of Formula I can befrom about 0.001% (w/v) to about 5% (w/v), from about 0.001% (w/v) toabout 1.0% (w/v), from about 0.01% (w/v) to about 0.5% (w/v), from about0.01% to about 1% (w/v), from about 0.1% to about 0.5% (w/v), or fromabout 0.5% to about 1% (w/v) in liquid and emulsion formulations. Theactual dose of the active compounds of the present invention depends onthe specific compound, and on the condition to be treated.

Emulsions may be prepared by combining a compound of Formula I in asterile lipophilic vehicle or fixed oil. The lipophilic vehicle or fixedoil may be selected from the group consisting of synthetic mono- anddiglycerides, fatty acids (including oleic acid), naturally occurringvegetable oils, sesame oil, coconut oil, peanut oil, cottonseed oil,castor oil, olive oil, mineral oil, synthetic fatty vehicles, and ethyloleate. Buffers, emulsifiers, dispersing agents, preservatives,antioxidants, and the like can be incorporated as required.

Pharmaceutically acceptable excipients for use with the invention mayinclude but are not limited to preservatives, buffering agents,antioxidants, lipophilic vehicles, hydrophilic vehicles, tonicityagents, electrolytes, thickeners, neutralizing agents, emulsifiers,dispersing agents, demulcents, plasticizers, occlusive agents, and filmformers, and combinations thereof. Certain compositions may include botha buffer component and a tonicity component.

Useful preservatives may include benzalkonium chloride, PURITE®, sodiumbisulfite, sodium bisulfate, sodium thiosulfate, ascorbate, benzalkoniumchloride, chlorobutanol, thimerosal, phenylmercuric acetate,phenylmercuric borate, phenylmercuric nitrate, methyl and ethylparabens, methylparaben, polyvinyl alcohol, benzyl alcohol,phenylethanol, hexetidine, chlorite components, such as stabilizedchlorine dioxide, metal chlorite, and other ophthalmically acceptablepreservatives. The concentration of the preservative component, if any,in the present compositions is a concentration effective to preserve thecomposition, and is often in a range of about 0.00001% to about 0.05% orabout 0.1% (w/w) of the composition.

Acceptable buffering agents may include HEPES and those prepared from asuitable combination of the acid and/or base forms of acetates,citrates, phosphates, carbonates, succinates, and borates, such assodium citrate dihydrate and boric acid. Phosphate buffers may becomposed of sodium phosphate dibasic and sodium phosphate monobasic.Examples include monosodium phosphate, monohydrate, sodium phosphatedibasic heptahydrate, and sodium phosphate monobasic monohydrate.Buffering agents may be provided in any of the compositions in an amounteffective to control the pH of the composition. The pH of thecomposition can be in a range of about 6 to about 8, about 7 to about 8,about 7 to about 7.6, or about 7.5 to about 8.

Useful tonicity agents may include glycerin, sugar alcohols, xylitol,sorbitol, glycerol, erythritol, mannitol, salts, potassium chlorideand/or sodium chloride. Tonicity agents may be provided in an amounteffective to control the tonicity or osmolality of the compositions. Theosmolality of the composition can be in a range of about 200 to about400, or about 250 to about 350, mOsmol/kg respectively. In oneembodiment, the composition is isotonic. An isotonic solution is asolution that has the same solute concentration as that inside normalcells of the body and the blood. An isotonic solution in contact with acell produces no net flow of water across the cell membrane. Usefullipophilic vehicles may include castor oil, squalane, diethylene glycolmonoethyl ether, propylene glycol, isostearyl isostearate, isopropylmyristate, dipropylene glycol dimethyl ether, diethylene glycol,dipropylene glycol, mineral oil, silicone oil, caprylic/caprictriglycerides, cetyl alcohols, and stearyl alcohols. Useful hydrophilicvehicles include water. A pharmaceutical composition may optionallycomprise an acceptable amount of dimethyl sulfoxide as an excipient.Additional examples of excipients that may be optionally included in thepharmaceutical compositions of the present invention may include thoselisted in Table A.

TABLE A Function Ingredient Active Compound of Formula I Thickener orcarbomer, sodium polyelectrolyte carboxymethylcellulose,methylcellulose, hydroxypropyl methylcellulose, polyvinyl alcohol,xanthan gum Neutralizing Agent sodium hydroxide, organic basesEmulsifier or polysorbate 20, polysorbate 40, dispersing agentpolysorbate 60, polysorbate 80, POE- 40-stearate, Pemulen ® and otherpolymeric emulsifiers. Demulcent carboxymethylcellulose sodium,hydroxypropyl methylcellulose hydroxyethyl cellulose, methylcellulose,polyvinyl alcohol, povidone, glycerin, propylene glycol, PEG 300, PEG400 Plasticizer Silicone oils, isostearyl alcohol, cetyl alcohol,glycerin Occlusive Agent silicone oils, petrolatum, waxes Film Formeracrylate/octylacrylamide copolymer, poly(ethyl acrylate, methylmethacrylate), chitosan, polyvinyl alcohol, polyisobutylene,polyvinylpyrrolidone-vinyl acetate copolymer, silicon gum,polyvinylpyrrolidone, other sustained release polymeric films

U.S. Pat. No. 5,474,979, the entire contents of which are incorporatedherein by reference, provides examples of emulsions that may be used toprepare pharmaceutical compositions of the present invention. The patentdiscloses the vehicle used in Restasis®, cyclosporin A 0.05%,manufactured by Allergan, Inc. This vehicle may be used to preparepharmaceutical compositions of the present invention.

Methods of Preparation

The present invention includes processes (i.e., methods) for preparingcompounds having Formula I. Compounds having Formula I may be preparedaccording to the following reaction schemes and accompanyingdiscussions. Unless otherwise indicated, the R¹, R², R³, R⁴, R⁵, R⁶, R⁷,R⁸, R⁹, R¹⁰, R¹², R¹³, and R¹⁴, and m, n, p, and q variables; andstructure of Formula I; in the following reaction schemes and discussionare as defined above in the Summary of the Invention.

The present invention includes isotopically-labeled compounds of FormulaI. For Example, a compound having Formula I may contain one or moreisotopic atoms such as deuterium ²H (or D) in place of proton ¹H (or H)or ¹³C in place of ¹²C and the like. Similar substitutions can beemployed for N, O and S. The use of isotopes may assist in analytical aswell as therapeutic aspects of the invention. For example, use ofdeuterium may increase the in vivo half-life by altering the metabolism(rate) of the compounds of the invention. These compounds can beprepared in accord with the preparations described by use ofisotopically enriched reagents.

Isotopically-labeled compounds of the present invention are identical tothose recited herein, except that one or more atoms in the compound arereplaced by an atom having an atomic mass or mass number different fromthe atomic mass or mass number usually found in nature. Examples ofisotopes that can be incorporated into compounds of the inventioninclude isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus,fluorine and chlorine and iodine, such as ²H, ³H, ¹¹C, ¹³C, ¹⁴C, ¹⁵N,¹⁸O, ¹⁷O, ³¹P, ³²P, ³⁵S, ¹⁸F, ³⁶Cl and ¹²³I, respectively.

The present invention can further include the Intermediates formed bythe following schemes.

As shown in the following schemes, the starting material for compoundsof Formula I is cyclosporin A (CAS Number 59865-13-3). Cyclosporin A maybe obtained commercially from suppliers such as Sigma-Aldrich (St.Louis, Mo., United States) or TCI America (Portland, Oreg., UnitedStates). Other cyclosporin starting materials such as Cyclosporin D (CASRegistry Number 63775-96-2) may also be obtained through commercialsuppliers such as Enzo Life Sciences (Ann Arbor, Mich., United States;Farmingdale, N.Y., United States). Other cyclosporin starting materialsmay be prepared from cyclosporin A as described by M. Mutter et al. Tet.Lett. 2000, 41, 7193-7196 and U.S. Pat. No. 5,214,130.

All the reagents, solvents, catalysts for which the synthesis is notdescribed are purchased from chemical vendors such as Sigma Aldrich,Fluka, Bio-Blocks, Combi-blocks, TCI, VWR, Lancaster, Oakwood, TransWorld Chemical, Alfa, Fisher, Maybridge, Frontier, Matrix, Ukrorgsynth,Toronto, Ryan Scientific, SiliCycle, Anaspec, Syn Chem, Chem-Impex,MIC-scientific, Ltd; however some known intermediates, were preparedaccording to published procedures.

In general, characterization of the compounds is performed according tothe following methods: Proton nuclear magnetic resonance (¹H NMR,written occasionally as ¹H NMR) and carbon nuclear magnetic resonance(¹³C NMR, written occasionally as ¹³C NMR) spectra were recorded on aBruker 300 or 500 MHz spectrometer in deuterated solvent. Chemicalshifts were reported as δ (delta) values in parts per million (ppm)relative to tetramethylsilane (TMS) as an internal standard (0.00 ppm)and multiplicities were reported as s, singlet; d, doublet; t, triplet;q, quartet; m, multiplet; bs, broad singlet; dd, doublet of doublets;and bt, broad triplet. Data were reported in the following format:chemical shift (multiplicity, integrated intensity, assignment).

Electron spray mass spectra (ESMS) were recorded on a Micromass ZQ.

The following abbreviations used in the following reaction schemes andaccompanying discussions are defined as follows:

-   -   Ac acetyl, a group of formula “—(C═O)CH₃”    -   DCM dichoromethane    -   CH₂Cl₂ dichloromethane    -   LDA lithium diisopropylamide    -   THF tetrahydrofuran    -   NMO 4-Methylmorpholine N-oxide    -   CH₃CN acetonitrile    -   TPAP Tetrapropylammonium perruthenate    -   MeOH methanol    -   NaCNBH₃ sodium cyanoborohydride    -   CD₃OD deuterated methanol    -   DMSO-d6 deuterated dimethyl sulfoxide    -   NaOMe sodium methoxide    -   EtOH ethanol    -   NaBH₄ sodium borohydride    -   MgSO₄ magnesium sulfate    -   NH₄Cl ammonium chloride    -   HCl hydrochloric acid    -   DIBAL-H Diisobutylaluminium hydride    -   Et₂O ether    -   K₂CO₃ potassium carbonate    -   DMF N,N-dimethylformamide    -   Et₃N triethylamine    -   CuI copper iodide    -   PdCl₂(PPh₃)₂ Bis(triphenylphosphine)palladium(II) chloride    -   NaH sodium hydride    -   EtOAc ethylacetate    -   AcOH acetic acid    -   TFA trifluoroacetic acid    -   NH₃ ammonia    -   CDCl₃ deuterated chloroform    -   n-Bu₄NOH Tetrabutylammonium hydroxide    -   NH₂NH₂ hydrazine    -   LAH or LiAlH₄ Lithium aluminium hydride    -   DEAD diethyl azodicarboxylate    -   Ph₃P triphenylphosphine    -   M molar concentration (molarity)    -   MPLC Medium pressure liquid chromatography    -   DIPEA diisopropylethylamine    -   i-Pr isopropyl    -   n-Bu n-butyl    -   TBDMSOTf t-Butyldimethylsilyl trifluoromethanesulphonate    -   OTBDMS O-t-Butyldimethylsilyl    -   TBDMS t-Butyldimethylsilyl    -   10% Pd/C 10% Palladium on carbon    -   NaBH(OAc)₃ or NaBHOAc3 Sodium triacetoxyborohydride    -   TBAF tetrabutylammonium fluoride    -   n-BuLi n-Butyl lithium    -   i-Pr₂NH Diisopropylamine    -   ESMS MH⁺ Electrospray mass spectrum positive ion    -   Grubbs II catalyst Grubbs catalyst second generation, also known        as        (1,3-Bis(2,4,6-trimethylphenyl)-2-imidazolidinylidene)dichloro        (phenylmethylene)(tricyclohexylphospine)ruthenium    -   HOBt-EDC N-hydroxybenzotriazole        1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride    -   Pd(PPh₃)₄ Tetrakis(triphenylphospine) palladium(0)    -   OTBDMS t-Butyldimethylsilyloxy    -   iBuOC(O)Cl Isobutyl chloroformate    -   MeSO2Cl Methane sulphonyl chloride

-   -   Pyridine    -   MeSSMe Dimethyl disulphide    -   DBU 1,8-Diazabicyclo[5.4.0]undec-7-ene    -   TBDMS t-Butyldimethylsilyl    -   H-cube Continuous flow hydrogenation apparatus    -   BuLi (same as n-BuLi above)    -   PhSSPh Diphenyl disulphide    -   DMAP 4-Dimethylaminopyridine    -   TEA Triethylamine    -   Hg(OAc)₂ Mercury(II) acetate    -   CSA Camphor sulphonic acid    -   MeI Methyl iodide    -   BnNEt3⁺Cl⁻ Benzytriethylammonium chloride    -   aq. KOH Aqueous potassium hydroxide    -   Ac2O Acetic anhydride    -   NB S N-Bromosuccinimide    -   AlBN 2,2′-Azobis(2-methylpropionitrile)    -   H2 or H₂ Hydrogen gas    -   10% Pd/C 10% palladium on carbon

-   -   -   dimethyl maleate

    -   O₃ ozone

-   -   -   morpholine

    -   ClCH₂CH₂Cl dichloroethane

    -   n-BuLi n-butyllithium

    -   i-Pr₂NH diisopropylamine

    -   CO₂ carbon dioxide

    -   ClCO₂CH₂Cl chloromethylchloroformate

    -   ClCO₂CH₂ CH₂Cl 2-chloroethylchloroformate

-   -   -   1-bromo-but-3-ene

    -   Cs₂CO₃ cesium carbonate

    -   LiOH lithium hydroxide

    -   Pd(PPh₃)₄ tetrakistriphenylphosphine palladium(0)

    -   PdCl₂(CH₃CN)₂ Bis(acetonitrile)dichloropalladium(II)

    -   ClCO₂CHMeCl 1-chloroethylchloroformate

    -   CCl₄ carbontetrachloride

-   -   -   3-[(N,N-diethyl)amino]propan-1-thiol

    -   Me methyl, a group of formula “—CH₃”

    -   Et ethyl, a group of formula “—CH₂CH₃”

    -   RT room temperature

    -   o/n overnight

The following synthetic schemes illustrate how compounds according tothe invention can be made. Those skilled in the art will be routinelyable to modify and/or adapt the following schemes to synthesize anycompound of the invention covered by Formula I.

Scheme I Procedure for Obtaining a Compound Having Formula I, Whereinn=0, m=0, and R⁸ is CH₂

Preparation of Intermediate 1

Intermediate 1, also known as [Methylene-Sar]³, is prepared as follows.

To a solution of diisopropylamine (11.2 ml, 80 mmol) in dry THF (240 ml)at −78° C. under an atmosphere of nitrogen was added dropwise n-butyllithium (2.5 M in hexane, 32 ml, 80 mmol) and the resulting mixture wasstirred at −78° C. for 60 minutes.

A solution of dry cyclosporine A (dried by azeotroping with 2×40 mltoluene then kept in dessicator o/n in presence of P₂O₅) (9.6 g, 8.0mmol) in dry THF (40 ml) was added and the reaction was stirred underthe same conditions for 2 h. A flow of carbon dioxide was bubbledthrough the reaction mixture for 30 minutes with temperature increasingto −50° C. The resulting mixture was allowed to warm to 15° C. over aperiod of 2 hours then cool back down to −50° C. before the addition ofchloromethylchloroformate (7.1 ml, 80 mmol). The reaction mixture wasallowed to warm to room temperature overnight then cooled to 0° C. andacetic acid (5 ml, 88 mmol) was added.

The mixture was allowed to warm to room temperature, the solventevaporated and the resultant mixture was partitioned between ethylacetate and brine. The organic phase was separated, dried (Na₂SO₄) andconcentrated to give a yellow oil.

The crude product was purified by MPLC chromatography using a solventgradient of 100% diethyl ether→96% diethyl ether/4% methanol to give[methylene-Sar]³cyclosporinA (Intermediate I).

ESMS MH⁺ 1214.8, MNa⁺ 1236.8

¹H NMR (CDCl₃, ppm) δ 4.98 (d, 1H, olefin CH₂), 5.25 (d, 1H, olefinCH₂), 7.17 (d, 1H, amide NH), 7.52 (d, 1H, amide NH), 7.59 (d, 1H, amideNH), 7.85 (d, 1H, amide NH).

¹³C NMR (CDCl₃, ppm) δ 143.96 (olefin C), 108.09 (olefin CH₂).

By substituting chloroethylchloroformate for chloromethylchloroformate(reactant 3 in Preparation of Intermediate 1), Intermediate 16 can beproduced (structure shown below). Intermediates 4 and 16 can serve asthe starting material for the production of amines and amides of FormulaI, as shown and described below.

Preparation of Intermediate 2

To a solution of [methylene-Sar]³cyclosporinA (5 g, 4.12 mmol)(Intermediate 1) in DMF (50 ml) at 0° C. under an atmosphere of nitrogenwas added triethylamine (10 eq., 5.75 ml, 41.2 mmol) followed by adropwise addition of TBDMSOTf (5 eq, 4.5 ml, 20.6 mmol) (over 5 minutes)and the reaction mixture warmed to room temperature over 2 h. Thereaction mixture was diluted with t-butyl methyl ether (200 ml), thenwashed with 2N HCl (100 ml), followed by H₂O (100 ml). The aqueousextracts were extracted with t-butyl methyl ether (100 ml), and thecombined organics washed with H₂O (2×100 ml), brine (100 ml) then dried(MgSO₄), filtered and evaporated under reduced pressure to yieldIntermediate 2 as a viscous oil. The crude product was used in the nextstep without further purification.

¹H NMR (CDCl₃, ppm) δ 7.40 (d, 1H, amide NH), 7.79 (d, 1H, amide NH),7.90 (d, 1H, amide NH), 8.28 (d, 1H, amide NH).

Preparation of Intermediate 3

A solution of[(5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-octanoicacid]¹[methylene-Sar]³cyclosporin A (Intermediate 2) (9 g, approx 4mmol) was dissolved in CH₂Cl₂ (200 ml) and added to a 3-neck flaskequipped with inlet (for nitrogen/ozone addition) and outlet connectedto a Dreschler bottle containing 2M KI solution. The reaction mixturewas cooled to −78° C. over a solid CO₂/acetone bath, under a nitrogenatmosphere. When the temperature of the reaction vessel had stabilised,the nitrogen was removed and ozone bubbled through the reaction mixtureuntil it became a pale blue colour (approx. 20 minutes). The ozonesupply was removed and nitrogen bubbled through the reaction mixtureuntil the blue colour had gone, then dimethylsulphide (0.8 ml) wasadded, and the reaction mixture warmed to room temperature over 2 hours.After this time, the reaction mixture was washed with H₂O (3×200 ml),then dried (MgSO₄), filtered and evaporated under reduced pressure toyield the crude product as a clear, viscous oil.

The crude product was purified by MPLC chromatography using a solventgradient of 100% hexane→40% ethyl acetate/60% hexane to giveIntermediate 3 as a white solid.

ESMS MH⁺ 1316.67

¹H NMR (CDCl₃, ppm) δ 7.53 (d, 1H, amide NH), 7.75 (d, 1H, amide NH),7.84 (d, 1H, amide NH), 8.33 (d, 1H, amide NH), 9.63 (s, 1H, aldehydeH).

Preparation of Intermediate 4

To a solution of[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-oxo-hexanoicacid]¹[methylene-Sar]³cyclosporin A (Intermediate 3) (1 g) in ethanolwas added 10% Palladium on Carbon (0.5 g) and the reaction stirred undera hydrogen atmosphere for 18 h. After this time, the reaction mixturewas filtered through a pad of celite and washed with ethyl acetate. Thesolvent was evaporated to leave the Intermediate 4 as a fluffy whitesolid. Obtain a mixture of (R) and (S)-methyl, approx >7:1 (R):(S). Theproduct can be used like this and purified later in the synthesis.

ESMS MH⁺ 1318.77

¹H NMR (CDCl₃, ppm) δ 7.57 (d, 1H, amide NH), 7.65 (d, 1H, amide NH),7.89 (d, 1H, amide NH), 8.47 (d, 1H, amide NH), 9.63 (s, 1H, aldehydeH).

Preparation of Intermediate 5

To a stirred solution of[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-oxo-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 4) (0.13 g, 0.1 mmol)in 1,2-dichloroethane (1 ml) was added morpholine (0.02 ml, 0.2 mmol)and sodium triacetoxyborohydride (0.038 g, 0.2 mmol) and the reactionmixture stirred at room temperature for 18 h. After this time, thereaction mixture was diluted with dichloromethane (5 ml) and washed withH₂O (3×5 ml), then the organic phase was dried (MgSO₄), filtered andevaporated. Intermediate 5 was obtained as a white solid and notpurified further at this stage.

ESMS MH⁺ 1389.78

¹H NMR (CDCl₃, ppm) δ 7.57 (d, 1H, amide NH), 7.64 (d, 1H, amide NH),7.96 (d, 1H, amide NH), 8.38 (d, 1H, amide NH).

Preparation of[(3R,4R,5S)-4-(hydroxy)-3-methyl-5-(methylamino)-1-(morpholin-4-yl)hexanoic acid]¹[(R)-Me-Sar]³cyclosporin A (Compound F)

To a stirred solution of[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Intermediate 5) (0.13 g, 0.1 mmol) inTHF (2 ml) was added tetrabutylammonium fluoride TBAF (1.0M soln in THF,0.2 ml, 0.2 mmol) and the reaction mixture stirred at room temperaturefor 8 h. After this time, the reaction solvent was evaporated, and theresulting residue redissolved in dichloromethane and washed with H₂O(3×5 ml). The organic phase was dried (MgSO₄), filtered and evaporatedto give a white solid. The crude product was first passed through a 5 gSCX (acidic) cartridge using methanol (collects TBDMS impurity) then 5%ammonia in methanol. The basic fractions were combined and evaporated togive[(3R,4R,5S)-4-(hydroxy)-3-methyl-5-(methylamino)-1-(morpholin-4-yl)hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Compound F) as a pure white solid.

ESMS MH⁺ 1275.69

¹H NMR (CDCl₃, ppm) δ 7.18 (d, 1H, amide NH), 7.39 (d, 1H, amide NH),7.76 (d, 1H, amide NH), 7.89 (d, 1H, amide NH).

While Scheme I shows morpholine as the reactant at Step 4, Intermediate4 can be reacted with other heterocycles or amines to produce a set ofcorresponding intermediates. Each of the resulting intermediates canthen be separately deprotected according to Step 5 of Scheme I toproduce a compound of Formula I. For instance, N-methylpiperazine,piperazinone, or diethylamine (NH(CH₂CH₃)₂) may be separately reactedwith Intermediate 4 to produce three distinct compounds of Formula Iwherein n=0 and m=0.

Preparation of Compound L

Thus, using the procedure shown in Scheme I with N-methylpiperazine asthe reactant at Step 4,[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-(4-methylpiperazino)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Intermediate 50) was obtained as awhite solid (ESMS MH⁺ 1402.81). Intermediate 50 was deprotected as shownin Step 5 to give Compound L([(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-(4-methylpiperazino)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A) as a white solid.

ESMS MH⁺ 1288.74

¹H NMR (CDCl₃, ppm) δ 7.20 (d, 1H, amide NH), 7.38 (d, 1H, amide NH),7.79 (d, 1H, amide NH), 7.89 (d, 1H, amide NH).

Preparation of Compound M

Using the procedure shown in Scheme I with diethylamine as the reactantat Step 4,[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-N-diethylamino-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Intermediate 51) was obtained as awhite solid (ESMS MH⁺ 1375.87).

Intermediate 51([(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-N-diethylamino-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A) was deprotected according to Step 5 inScheme I to give Compound M([(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-diethylamino-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A) as a white solid.

ESMS MH⁺ 1261.82

¹H NMR (CDCl₃, ppm) δ 7.18 (d, 1H, amide NH), 7.39 (d, 1H, amide NH),7.80 (d, 1H, amide NH), 7.82 (d, 1H, amide NH).

Preparation of Compound N

Using the procedure shown in Scheme I with piperazinone as the reactantat Step 4,[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-(N-3-piperazinone)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Intermediate 52) was obtained as awhite solid (ESMS MH⁺ 1402.88).

Intermediate 52([(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-(N-3-piperazinone)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A) was then deprotected according to Step5 in Scheme I to give Compound N([(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-(N-3-piperazinone)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A), as a white solid.

ESMS MH⁺ 1288.81

¹H NMR (CDCl₃, ppm) δ 7.12 (d, 1H, amide NH), 7.91 (d, 1H, amide NH),8.10 (d, 1H, amide NH), 8.32 (d, 1H, amide NH).

Using the procedure shown in Scheme I with the appropriate amines thefollowing compounds were prepared:

Compound AK (see Table 17)[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-{1H-imidazol-4-yl}-ethylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1300.4

¹H NMR (CDCl₃, ppm) δ 6.79 (bs, 1H, imidazole CH), 7.25 (d, 1H, amideNH), 7.54 (bs, 2H, imidazole CH and amide NH), 7.82 (d, 1H, amide NH),8.04 (d, 1H, amide NH).

Compound AF (see Table 16)[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-methoxyethylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1263.6

¹H NMR (CDCl₃, ppm) δ 7.20 (d, 1H, amide NH), 7.40 (d, 1H, amide NH),7.80 (d, 1H, amide NH), 7.89 (d, 1H, amide NH).

Compound AG (see Table 16)[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-methoxyethyl)methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1277.6

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.41 (d, 1H, amide NH),7.78 (d, 1H, amide NH), 7.89 (d, 1H, amide NH).

Compound O (see Table 9),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(N-(3aR*,6aS*)-2-methyloctahydropyrrolo[3,4-c]pyrrolo)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1314.7

¹H NMR (CDCl₃, ppm) δ 7.21 (d, 1H, amide NH), 7.41 (d, 1H, amide NH),7.79 (d, 1H, amide NH), 7.79 (d, 1H, amide NH).

Compound AJ (see Table 16),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(1,4-dioxan-2-ylmethyl)methylamino-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1319.6

¹H NMR (CDCl₃, ppm) δ 7.13 (d, 1H, amide NH), 7.44 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 7.91 (d, 1H, amide NH).

Compound J (see Table 10),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(thiomorpholino)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A

ESMS MH⁺ 1291.58

¹H NMR (CDCl₃, ppm) δ 7.18 (d, 1H, amide NH), 7.42 (d, 1H, amide NH),7.76 (d, 1H, amide NH), 7.92 (d, 1H, amide NH).

Compound P (see Table 11),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(1,1-dioxo-thiomorpholino)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A

ESMS MH⁺ 1323.77

¹H NMR (CDCl₃, ppm) δ 7.14 (d, 1H, amide NH), 7.51 (d, 1H, amide NH),7.71 (d, 1H, amide NH), 7.97 (d, 1H, amide NH).

Compound X (see Table 13),[[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-homomorpholino-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1289.67

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.41 (d, 1H, amide NH),7.78 (d, 1H, amide NH), 7.91 (d, 1H, amide NH).

Compound EK (see Table 13),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2,2,6,6-tetrafluoro-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1347.67

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.53 (d, 1H, amide NH),7.73 (d, 1H, amide NH), 8.08 (d, 1H, amide NH).

Compound EL[(3R,4R,5S)-(3,3-Difluoro-pyrrolidin-1-yl)-4-Hydroxy-3-methyl-5-(methylamino)-1-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1295.91

¹H NMR (CDCl₃, ppm) δ 7.17 (d, 1H, amide NH), 7.42 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 7.92 (d, 1H, amide NH).

Compound EM[(3R,4R,5S)-(3,3-Difluoro-azetidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-1-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1281.56

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.41 (d, 1H, amide NH),7.76 (d, 1H, amide NH), 7.92 (d, 1H, amide NH).

Compound EN[(3R,4R,5S)-(4,4-Difluoro-piperidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-1-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1309.55

¹H NMR (CDCl₃, ppm) δ 7.16 (d, 1H, amide NH), 7.42 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 7.90 (d, 1H, amide NH).

Compound ER[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(3,3,4,4-tetrafluoro-pyrrolidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1331.52

¹H NMR (CDCl₃, ppm) δ 7.16 (d, 1H, amide NH), 7.50 (d, 1H, amide NH),7.73 (d, 1H, amide NH), 8.03 (d, 1H, amide NH).

Compound EQ(3R,4R,5S)-1-[Bis-(3,3,3-trifluoro-propyl)-amino]-[(4-Hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ES/MS: 1398.0 MH⁺

¹H NMR (CDCl₃, ppm) δ 7.11 (d, 1H, amide NH), 7.52 (d, 1H, amide NH),7.73 (d, 1H, amide NH), 7.98 (d, 1H, amide NH).

Scheme II Procedure for Obtaining a Compound Having Formula I, Whereinn=3 and m=0

The distance between the amine in the side chain at position 1 and thecyclosporin scaffold can be varied (that is, the value of n can beadjusted between 0 and 4) according to the schemes set forth herein.

For example Scheme II enables the practitioner to obtain a compound ofFormula I having an amine at the position 1 α-carbon in which n=3.

Preparation of Intermediate 6

[Methylene-Sar]³cyclosporin A (Intermediate 1) (242 mg, 0.2 mmol) and4-bromo but-1-ene, (405 mg, 3.0 mmol) were dissolved in drydichloromethane (4 mL). The mixture was purged with nitrogen before theaddition of Grubbs catalyst second generation (40 mg, 0.048 mmol) thenrefluxed for 18 hours. The reaction mixture was cooled to roomtemperature then purified by silicagel chromatography using a solventgradient of 100% dichloromethane→96% dichloromethane/4% methanol toprovide[(3E,6R,7R,8S)-1-bromo-7-hydroxy-6-methyl-8-(methylamino)-non-3-enoicacid]¹cyclosporin A (Intermediate 6) as an off-white solid.

Grubbs' catalyst second generation (Grubbs II catalyst) is described inU.S. Patent Application Publication No. 2003/0186855. Longer chainbrominated alkenes may be used at Step 1 if desired to produce a longerside chain at position 1.

ESMS MH⁺ 1306.7/1308.7

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.65 (d, 1H, amide NH),7.70 (d, 1H, amide NH), 8.07 (d, 1H, amide NH).

Preparation of Intermediate 25

3E,6R,7R,8S)-1-bromo-7-hydroxy-6-methyl-8-(methylamino)-non-3-enoicacid]¹cyclosporin A (Intermediate 6) (260 mg, 0.2 mmol) was dissolved inacetonitrile. Morpholine (385 uL, 0.4 mmol), cesium carbonate (255 mg,0.8 mmol) and a catalytic amount of potassium iodide were added. Thereaction mixture was stirred at room temperature for 18 hours. Furtheramount of morpholine (100 uL, 0.1 mmol) was added and the reactionmixture stirred for 3 hours before evaporating the volatiles. Theresidue was purified by SCX chromatography using a solvent gradient of100% methanol→0.21M ammonia in methanol. The material obtained wasdissolved in ethyl acetate, washed three times with water, dried oversodium sulphate and concentrated to give[(3E,6R,7R,8S)-7-hydroxy-6-methyl-8-(methylamino)-1-N-morpholino-non-3-enoicacid]¹[methylene-Sar]³cyclosporin A (Intermediate 25) as a white solid.

ESMS MH⁺ 1313.8

¹H NMR (CDCl₃, ppm) δ 7.17 (d, 1H, amide NH), 7.62 (m, 2H, amide NH),8.00 (d, 1H, amide NH).

Preparation of Compound G

[(3E,6R,7R,8S)-7-hydroxy-6-methyl-8-(methylamino)-1-N-morpholino-non-3-enoicacid]¹[methylene-Sar]³cyclosporin A (Intermediate 25) (68 mg, 0.054mmol) was dissolved in ethanol (6 mL), treated with 10% palladium oncarbon (68 mg) then placed under a 1 atm atmosphere of hydrogen for 65hours. Fresh catalyst (68 mg) was added to the reaction mixture andhydrogenation carried on for an additional 24 hours. The reactionmixture was filtered through celite then concentrated in vacuo. Theresidue was purified by silicagel chromatography using a solventgradient of 100% dichloromethane→95% dichloromethane/5% methanol toprovide[(6R,7R,8S)-7-hydroxy-6-methyl-8-(methylamino)-1-N-morpholino-nonanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound G) as a white solid.

ESMS MH⁺ 1317.8

¹H NMR (CDCl₃, ppm) δ 7.17 (d, 1H, amide NH), 7.50 (d, 1H, amide NH),7.69 (d, 1H, amide NH), 7.91 (d, 1H, amide NH).

Using the procedure shown in Scheme II with the appropriate amines thefollowing compounds were prepared:

Compound AH (see Table 16)[(6R,7R,8S)-7-Hydroxy-6-methyl-8-(methylamino)-1-N-diethylamino-nonanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1303.8

¹H NMR (CDCl₃, ppm) δ 7.14 (d, 1H, amide NH), 7.50 (d, 1H, amide NH),7.67 (d, 1H, amide NH), 7.93 (d, 1H, amide NH).

Compound AI (see Table 16)[(6R,7R,8S)-7-Hydroxy-6-methyl-8-(methylamino)-1-N-(2-methoxy)ethylamino-nonanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1305.7

¹H NMR (CDCl₃, ppm) δ 7.16 (d, 1H, amide NH), 7.51 (d, 1H, amide NH),7.67 (d, 1H, amide NH), 7.93 (d, 1H, amide NH).

Compound C (see Table 4)[(6R,7R,8S)-7-Hydroxy-6-methyl-8-(methylamino)-1-N-morpholinyl-nonanoicacid]¹cyclosporin A

ESMS MH⁺ 1303.8

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.5 (d, 1H, amide NH), 7.7(d, 1H, amide NH), 7.95 (d, 1H, amide NH).

Scheme III Procedure for Obtaining a Compound Having Formula I wheren=1, and m=1

Schemes III and IV describe efficient processes for the synthesis ofamides of Formula I. Scheme III describes the use of metathesis usingGrubbs II catalyst while Scheme IV describes the use of Wittigchemistry. The metathesis route (scheme III) is two steps shorter thanthe Wittig route (scheme IV) starting from intermediate 1.

Preparation of Intermediate 7

Methylene-Sar]³cyclosporin A (Intermediate 1) (242 mg, 0.2 mmol) anddimethyl maleate (376 uL, 3.0 mmol) were dissolved in drydichloromethane (4 mL). The mixture was purged with nitrogen before theaddition of Grubbs catalyst second generation (40 mg, 0.048 mmol) thenrefluxed for 18 hours. The reaction mixture was cooled to roomtemperature then purified by silicagel chromatography using a solventgradient of 100% dichloromethane→96% dichloromethane/4% methanol toprovide [[2E,5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-oct-2-enedioicacid]methyl ester]¹[methylene-Sar]³cyclosporin A (Intermediate 7) as anoff-white solid.

ESMS MH⁺ 1258.98

¹H NMR (CDCl₃, ppm) δ 5.74 (d, 1H, olefin), 7.00 (dt, 1H, olefin), 7.18(d, 1H, amide NH), 7.51 (d, 1H, amide NH), 7.60 (d, 1H, amide NH), 7.79(d, 1H, amide NH).

Preparation of Intermediate 8

[[(2E,5R,6R,7S)-6-Hydroxy-5-methyl-7-(methylamino)-oct-2-enedioicacid]methyl ester]¹[methylene-Sar]³cyclosporin A (Intermediate 7) (68mg, 0.054 mmol) was dissolved in ethanol (6 mL), treated with 10%palladium on carbon (68 mg) then placed under a 1 atm atmosphere ofhydrogen for 65 hours. Fresh catalyst (68 mg) was added to the reactionmixture and hydrogenation carried on for an additional 24 hours. Thereaction mixture was filtered through celite then concentrated in vacuo.The residue was purified by silicagel chromatography using a solventgradient of 100% dichloromethane→95% dichloromethane/5% methanol toprovide [[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-octanedioicacid]methyl ester]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 8) as awhite solid.

ESMS MH⁺ 1262.71

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.48 (d, 1H, amide NH),7.70 (d, 1H, amide NH), 7.78 (d, 1H, amide NH).

Preparation of Intermediate 9 from Intermediate 8

[[(5R,6R,7S)-6-Hydroxy-5-methyl-7-(methylamino)-octanedioic acid]methylester]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 8) (0.34 g, 0.27mmol) was dissolved in tetrahydrofuran (30 mL) then treated with asolution of lithium hydroxide (64 mg, 1.5 mmol) in water (4 mL). Thereaction mixture was stirred at room temperature for 25 hours thenconcentrated. The residue was treated with ethyl acetate and HCl 1N. Theorganic phase was washed with brine, dried (Na₂SO₄) and concentrated.The residue was purified by silicagel chromatography using a solventgradient of 100% dichloromethane→95% dichloromethane/5% methanol toprovide [(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-octanedioicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 9) as a white solid.

ESMS MH⁺ 1248.6

¹H NMR (CDCl₃, ppm) δ 7.21 (d, 1H, amide NH), 7.61 (d, 1H, amide NH),7.69 (d, 1H, amide NH), 8.03 (d, 1H, amide NH).

As described below, the procedure shown in Scheme III can be generallyadapted to produce a variety of Compounds of Formula I, depending on thestructure of the amine or heterocycle reacted with Intermediate 9 atStep 4 in Scheme III.

Preparation of Compound H from Intermediate 9

[(5R,6R,7S)-6-Hydroxy-5-methyl-7-(methylamino)-octanedioicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 9) (50 mg, 0.04 mmol)was dissolved in acetonitrile (2 mL) and stirred at 0° C. undernitrogen. Morpholine (8 uL, 0.09 mmol) then N-hydroxybenzotriazole (10mg, 0.05 mmol) then 1-ethyl-3-(3-dimethylaminopropyl) carbodiimidehydrochloride (11 mg, 0.056 mmol) were added. The reaction mixture wasleft to stir at 0° C. for 30 minutes then allowed to warm up to roomtemperature and stirred for 17 hours. The volatiles were removed and theresidue was partitioned between ethyl acetate and water, the organicphase was washed with water, brine then dried over sodium sulphate thenconcentrated in vacuo to give[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-N-morpholino-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound H) as a white solid.

ESMS MH⁺ 1318.0

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.52 (d, 1H, amide NH),7.72 (d, 1H, amide NH), 8.01 (d, 1H, amide NH).

Preparation of Compound T from Intermediate 9

By substituting N-methyl piperazine for morpholine in the proceduredescribed above to prepare Compound H,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(4-methylpiperazin-1-yl)-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound T) was obtained as awhite solid after purification by SCX chromatography using a solventgradient of 100% methanol→0.14M ammonia in methanol.

ESMS MH⁺ 1330.8

¹H NMR (CDCl₃, ppm) δ 7.14 (d, 1H, amide NH), 7.51 (d, 1H, amide NH),7.71 (d, 1H, amide NH), 8.00 (d, 1H, amide NH).

Preparation of Compound U from Intermediate 9

By substituting diethylamine for morpholine in the procedure describedabove to prepare Compound H,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-diethylamino-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound U) was obtained as awhite solid after purification by silicagel chromatography using asolvent gradient of 100% dichloromethane→95% dichloromethane/5%methanol.

ESMS MH⁺ 1303.8

¹H NMR (CDCl₃, ppm) δ 7.12 (d, 1H, amide NH), 7.50 (d, 1H, amide NH),7.69 (d, 1H, amide NH), 7.98 (d, 1H, amide NH).

Preparation of Compound W from Intermediate 9

By substituting 2-amino-ethanesulfonic acid dimethylamide for morpholinein the procedure described above to prepare Compound H,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-{sulfonic aciddimethylamide}-ethylamino)-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound W) was obtained as awhite solid after purification by SCX chromatography using methanol.

ESMS MH⁺ 1382.6

¹H NMR (CDCl₃, ppm) δ 6.99 (t, 1H, amide NH), 7.23 (d, 1H, amide NH),7.62 (d, 1H, amide NH), 7.80 (d, 1H, amide NH), 8.12 (d, 1H, amide NH).

Preparation of Compound Y from Intermediate 9

By substituting 2-(1H-imidazol-4-yl)-ethylamine for morpholine in theprocedure described above to prepare Compound H,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-{1H-imidazol-4-yl}-ethylamino)-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound Y) was obtained as awhite solid after purification by SCX chromatography using a solventgradient of 100% methanol→0.14M ammonia in methanol.

ESMS MH⁺ 1341.9

¹H NMR (CDCl₃, ppm) δ 6.61 (m, 1H, amide NH), 6.82 (bs, 1H, imidazoleCH), 7.22 (d, 1H, amide NH), 7.56 (bs, 1H, imidazole CH), 7.64 (d, 1H,amide NH), 7.78 (d, 1H, amide NH), 8.12 (d, 1H, amide NH).

Preparation of Compound Z from Intermediate 9

By substituting thiomorpholine 1,1-dioxide for morpholine in theprocedure described above to prepare Compound H,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-({1,1-dioxo}thiomorpholin-4-yl)-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound Z) was obtained as awhite solid after purification by SCX chromatography using methanol.

ESMS MH⁺ 1365.3

¹H NMR (CDCl₃, ppm) δ 7.21 (d, 1H, amide NH), 7.58 (d, 1H, amide NH),7.74 (d, 1H, amide NH), 8.08 (d, 1H, amide NH).

-   -   The chemical structures, and therefore formulas, of Compounds H,        T, U, W, Y, and Z are shown below.

Preparation of Compound ZZ from Intermediate 9

By substituting (3aR*,6aS*)-2-methyloctahydropyrrolo[3,4-c]pyrrole formorpholine in the procedure described above to prepare Compound H,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(N-(3aR*,6aS*)-2-methyloctahydropyrrolo[3,4-c]pyrrolo)-1-oxo-octanoicacid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZZ) was obtained as awhite solid after purification by SCX chromatography using a solventgradient of 100% methanol to 0.14M ammonia in methanol.

ESMS MH+1356.6

1H NMR (CDCl₃, ppm) δ 7.13 (d, 1H, amide NH), 7.50 (d, 1H, amide NH),7.71 (d, 1H, amide NH), 8.00 (d, 1H, amide NH).

Preparation of Compound ZY from Intermediate 9

By substituting 2-methoxy-ethylamine for morpholine in the proceduredescribed above to prepare Compound H,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-methoxyethylamino)-1-oxo-octanoicacid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZY) was obtained as awhite solid after purification by silica gel chromatography using asolvent gradient of 100% dichloromethane to 95% dichloromethane/5%methanol.

ESMS MH+1306.0

1H NMR (CDCl₃, ppm) δ 6.39 (bt, 1H, amide NH), 7.20 (d, 1H, amide NH),7.59 (d, 1H, amide NH), 7.78 (d, 1H, amide NH), 8.11 (d, 1H, amide NH).

Preparation of Compound ZX from Intermediate 9

By substituting (1,4-dioxan-2-ylmethyl)amine for morpholine in theprocedure described above to prepare Compound H,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(1,4-dioxan-2-ylmethyl)amino)-1-oxo-octanoicacid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZX) was obtained as awhite solid after purification by silica gel chromatography using asolvent gradient of 100% dichloromethane to 95% dichloromethane/5%methanol.

ESMS MH+1347.7

1H NMR (CDCl₃, ppm) δ 6.41 (dd, 1H, amide NH), 7.20 (d, 1H, amide NH),7.62 (bs, 2H, amide NH), 7.78 (d, 1H, amide NH), 8.12 (d, 1H, amide NH).

Using the procedure described in scheme III with piperidine in step 4,compound Q was prepared.

Compound Q (see Table 11)[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-piperidino-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A

ESMS MH⁺ 1315.6

¹H NMR (CDCl₃, ppm) δ 7.14 (d, 1H, amide NH), 7.51 (d, 1H, amide NH),7.71 (d, 1H, amide NH), 8.02 (d, 1H, amide NH).

Using the procedure described in scheme III with Cyclosporin A as thestarting material and morpholine as the amine in step 4, compound E wasprepared.

Compound E[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-N-morpholino-1-oxo-octanoicacid]¹cyclosporin A

ESMS MH⁺ 1303.68

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.5 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 8.1 (d, 1H, amide NH).

Scheme IV Procedure for Obtaining a Compound Having Formula I where n=1and m=1

For Scheme IV, Intermediate 4 is prepared as described above in SchemeI.

Preparation of Intermediate 10 from Intermediate 4

To a suspension of lithium chloride (127 mg, 3 mmol) in acetonitrile (5mL) was added successively diisopropylethylamine (522 uL, 3 mmol) andallyldiethylphosphono acetate (89 uL, 0.42 mmol). The mixture wasstirred at room temperature for 15 minutes before the addition of asolution of[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-oxo-hexanoicacid]1[(R)-methyl-Sar]3cyclosporin A (Intermediate 4) (393 mg, 0.3 mmol)in acetonitrile/methanol (50/50, 8 mL). The reaction mixture was stirredat room temperature for 18 hours then concentrated in vacuo. The residuewas partitioned between ethyl acetate and 1M HCl, the organic phase waswashed twice with further 1M HCl then saturated aqueous sodiumbicarbonate. The organic phase was dried over sodium sulphate thenconcentrated in vacuo. The residue was purified by silica gelchromatography using a solvent gradient of 100% dichloromethane to 95%dichloromethane/5% methanol to provide[(2E,5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-oct-2-enedioicacid]allyl ester]1[(R)-methyl-Sar]3cyclosporin A (Intermediate 10) as awhite solid.

1H NMR (CDCl₃, ppm) δ 5.73 (d, 1H, olefin), 7.06 (dt, 1H, olefin), 7.5(m, 2H, amide NH), 7.90 (d, 1H, amide NH), 8.37 (d, 1H, amide NH).

Preparation of Intermediate 11 from Intermediate 10

A solution of[[(2E,5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-octanedioicacid]allyl ester]1[(R)-methyl-Sar]3cyclosporin A (Intermediate 10) (97mg, 0.069 mmol) in dry THF (4 mL) had nitrogen gas bubbled through for 5minutes before and after the addition oftetrakis(triphenylphosphine)palladium (0) (16 mg, 0.0138 mmol).Morpholine (62 uL, 0.69 mmol) was then added and the reaction mixturewas stirred at room temperature for 20 hours. The volatiles were removedand the residue was dissolved in ethyl acetate and washed twice with 1MHCl then once with brine. The organic phase was dried over sodiumsulphate then concentrated in vacuo. NMR analysis indicated that thereaction was not complete and so the mixture was treated again asdescribed above and this time stirred at room temperature for 3 days.The above work-up provided a residue which was purified by silica gelchromatography using a solvent gradient of 100% dichloromethane to 96%dichloromethane/4% methanol to provide[(2E,5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-oct-2-enedioicacid]1[(R)-methyl-Sar]3cyclosporin A (Intermediate 11) as a white solid.

1H NMR (CDCl₃, ppm) δ 7.49 (d, 1H, amide NH), 7.59 (d, 1H, amide NH),7.78 (d, 1H, amide NH), 8.42 (d, 1H, amide NH).

Preparation of Intermediate 12 from Intermediate 11

A solution of[(2E,5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-oct-2-enedioicacid]1[(R)-methyl-Sar]3cyclosporin A (Intermediate 11) (56 mg, 0.041mmol) in dry acetone (2 mL) had nitrogen gas bubbled through it for 5minutes before the addition of bis(acetonitrile)dichloropalladium (II)(1 mg, 0.004 mmol). The reaction mixture was stirred at room temperaturefor 17 hours, then additional dry acetone (2 mL) andbis(acetonitrile)dichloropalladium (II) (1 mg, 0.004 mmol) were addedand the reaction mixture was stirred for a further 3 days. The reactionmixture was filtered through celite using diethyl ether and methanol assolvents. The solvents were evaporated and the residue obtained waspurified by silica gel chromatography using a solvent gradient of 100%dichloromethane to 95% dichloromethane/5% methanol to provide[(2E,5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-oct-2-enedioicacid]1[(R)-methyl-Sar]3cyclosporin A (Intermediate 12) as a white solid.

ESMS MH+1246.7

1H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.50 (d, 1H, amide NH),7.65 (d, 1H, amide NH), 8.05 (d, 1H, amide NH).

Preparation of Intermediate 9 from Intermediate 11

[(2E,5R,6R,7S)-6-(t-Butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-oct-2-enedioicacid]1[(R)-methyl-Sar]3cyclosporin A (Intermediate 11) (100 mg, 0.073mmol) was dissolved in methanol (20 mL), treated with 10% palladium oncarbon (50 mg) then placed under a 1 atm atmosphere of hydrogen for 17hours. The reaction mixture was filtered through celite thenconcentrated in vacuo to provide the dihydro version of intermediate 11[[(5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-octanedioicacid]1[(R)-methyl-Sar]3 cyclosporin A (Intermediate 56) as a whitesolid.

1H NMR (CDCl₃, ppm) δ 7.49 (d, 1H, amide NH), 7.83 (m, 2H, amide NH),8.30 (d, 1H, amide NH).

A solution of[(5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-octanedioicacid]1[(R)-methyl-Sar]3cyclosporin A (Intermediate 56) (95 mg, 0.07mmol) in tetrahydrofuran (4 mL) was treated with a 1M solution oftetrabutylammonium fluoride in tetrahydrofuran (350 uL, 0.35 mmol). Thereaction mixture was stirred at room temperature for 23 hours. Thevolatiles were removed and the residue was partitioned between ethylacetate and water, the organic phase was washed twice with water thendried over sodium sulphate then concentrated in vacuo. The residue waspurified by silicagel chromatography using a solvent gradient of 100%dichloromethane to 94% dichloromethane/6% methanol to provide

[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-octanedioicacid]1[(R)-methyl-Sar]3 cyclosporin A (Intermediate 9) as a white solid.

ESMS MH+1248.6

1H NMR (CDCl₃, ppm) δ 7.21 (d, 1H, amide NH), 7.61 (d, 1H, amide NH),7.69 (d, 1H, amide NH), 8.03 (d, 1H, amide NH).

The final step in Scheme IV, the conversion of Intermediate 9 to aCompound of Formula I, is carried out as described above in Scheme III,step 4.

Scheme V Procedure for Obtaining a Compound Having Formula I where n=2and m=0

Preparation of Intermediate 9

For purposes of Scheme V, Intermediate 9 is prepared as described inScheme IV.

Preparation of Intermediate 13

To a solution of Intermediate 9 in a solvent such as dichloromethane isadded isobutyl carbonyl chloride together with a base such astriethylamine. After removal of the solid by filtration the solvent isevaporated to give the anhydride. The anhydride is dissolved in asolvent such as dioxane and a reducing agent such as sodium borohydrideis added in order to give the alcohol, intermediate 13.

Preparation of a Compound of Formula I from Intermediate 13

Intermediate 13 is dissolved in DCM and methanesulphonyl chloride andpyridine are added. The product is dissolved in a solvent such asacetonitrile and a base such as cesium carbonate is added. The requiredamine is added and the reaction mixture is stirred at a temperaturebetween room temperature and 60° in order to give compounds of Formula Ias shown in scheme V.

By using an alternative starting aldehyde, such as Intermediate 15,shown below, containing one less carbon and using the same chemistrydescribed in Schemes IV and Xa, amines are prepared where n=1 and m=0,and amides are prepared where n=0 and m=1. The alternative startingaldehyde used for such syntheses can be prepared as shown below inScheme VI.

Scheme VI Preparation of Intermediate 15 (aldehyde)

Preparation of Intermediate 4

For purposes of Scheme VI, Intermediate 4 is prepared according toScheme I.

Preparation of Intermediate 14

To a solution of[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-oxo-hexanoicacid]¹[methylene-Sar]³cyclosporin A (Intermediate 4) (0.4 g) in DMF (1ml) cooled to 0° C. was added DBU (0.5 ml) followed by a dropwisesolution of TBDMSOTf (0.38 ml) in DMF (0.5 ml) and the reaction stirredunder a nitrogen atmosphere for 18 h. After this time, the reactionmixture was diluted with ethyl acetate (10 ml) and H₂O (10 ml). Theorganic layer was separated, dried, filtered and evaporated underreduced pressure.

The crude product was purified by MPLC chromatography using a solventgradient of 100% dichloromethane→2% methanol/98% dichloromethane to giveIntermediate 14 as a white solid.

¹H NMR (CDCl₃, ppm) 7.43 (d, 1H, amide NH), 7.58 (d, 1H, amide NH), 7.93(d, 1H, amide NH), 8.58 (d, 1H, amide NH).

Preparation of Intermediate 15

A solution of Intermediate 14 (0.33 g, approx 0.23 mmol) was preparedaccording to the method used for the preparation of intermediate 3(scheme 1, step 2) to give[(2R,3R,4S)-3-(t-butyldimethylsilanyloxy)-2-methyl-4-(methylamino)-1-oxo-pentanoicacid]¹[methylene-Sar]³cyclosporin A, intermediate 15 as a clear oil.

¹H NMR (CDCl₃, ppm) 7.44 (d, 1H, amide NH), 7.47 (d, 1H, amide NH), 7.97(d, 1H, amide NH), 8.67 (d, 1H, amide NH), 9.52 (s, 1H, aldehyde H).

Scheme VII Procedure for Obtaining a Compound Having Formula I where R⁸is CH₂O, n=2, and m=0

Preparation of Intermediate 53

Intermediate 53 was prepared by reduction of the aldehyde, Intermediate4, using sodium borohydride in a solvent such as methanol, as describedin US 2004/0110666.

Specifically, Intermediate 4 (0.17 g, 0.13 mmoles) was dissolved inmethanol (5 ml) and sodium borohydride (0.01 g, 0.26 mmoles) was addedwith stirring. After stirring for 3 hours the reaction mixture wasevaporated and the residue was partitioned between ethyl acetate andwater. The ethyl acetate layer was separated and dried over anhydrousmagnesium sulphate. The product was evaporated to give Intermediate 53.

1H NMR (CDCl₃, ppm) δ 7.57 (d, 1H, amide NH), 7.81 (d, 1H, amide NH),7.72 (d, 1H, amide NH), 8.40 (d, 1H, amide NH).

Preparation of Intermediate 54

Intermediate 53 (0.18 g, 0.13 mmoles was dissolved in THF (2 ml) anddeprotected by stirring overnight with 1M tetrabutylammonium fluoride(0.2 ml, 0.2 mmoles). The solvent was evaporated and the residue wastaken up in dichloromethane, washed with water (×3) and dried overanhydrous magnesium sulphate. The solvent was removed by evaporation togive crude compound which was purified by MPLC to give the requiredIntermediate 54.

1H NMR (CDCl₃, ppm) δ 7.32 (d, 1H, amide NH), 7.53 (d, 1H, amide NH),7.86 (d, 1H, amide NH), 8.17 (d, 1H, amide NH).

Preparation of Compound of Formula I from Intermediate 54

Intermediate 54 (0.07 g, 0.058 mmoles) was dissolved in DCM (1 ml) and2-bromo-N,N-diethylethylamine hydrobromide (0.02 g, 0.058 mmoles) wasadded to the reaction mixture followed by 40% aqueous potassiumhydroxide (1 ml) and the reaction mixture was stirred at roomtemperature overnight. The reaction mixture was diluted with water (15mL) and more DCM (15 mL) was added then the DCM layer was separated anddried over anhydrous magnesium sulphate. The product was evaporated andthe residue was purified using preparative thin layer chromatography(×2) to give a compound having Formula I (in this instance,[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-2-(N,N-diethylamino)ethoxy-hexanoicacid]¹[(S)-thio-isopropyl-Sar]³cyclosporin A (Compound AL)). Other alkylhalides such as iodomethane and 4-(2-chloroethyl) morpholine may be usedin a similar manner to prepare other compounds of Formula I according toScheme VII.

Scheme VIII

Procedure for obtaining a compound having Formula I, wherein R¹ is—CH₂CH₃, R¹¹ is O, n=0, m=0, and p=1. Scheme VIII:

Preparation of Intermediate 26 (Step 1)

Intermediate 26 is prepared according to the procedure used to prepareIntermediate 1 except that chloroethyl-chloro formate is used instead ofchloromethly-chloro formate.

Preparation of Intermediate 16 (Step 2)

Step 2 in Scheme VIII above was carried out in an identical manner asdescribed in step 2 Scheme 1 to give intermediate 16. Intermediate 16may be substituted for Intermediate 1 in any of the schemes shown aboveto obtain compounds of Formula I in which R¹ is ethyl.

Preparation of Intermediate 27 (Step 3)

To a stirred solution of[(3R,4R,5S)-4-(hydroxy)-3-methyl-5-(methylamino)-1-oxo-hexanoicacid]¹[ethyl-2-ene-Sar]³cyclosporin A (Intermediate 16) (0.219 g, 0.18mmol) in dichloromethane (10 ml) was added morpholine (0.079 ml, 0.9mmol) and sodium triacetoxyborohydride (0.160 g, 0.9 mmol) and thereaction mixture stirred at room temperature for 18 h. After this time,additional amounts of morpholine (0.079 ml, 0.9 mmol) and sodiumtriacetoxyborohydride (0.160 g, 0.9 mmol) were added and the reactionmixture stirred at 40° C. for 4.5 h. After this time, additional amountsof morpholine (0.025 ml, 0.28 mmol) and sodium triacetoxyborohydride(0.035 g, 0.2 mmol) were added and the reaction mixture stirred at 40°C. for 66 h. The reaction mixture was diluted with dichloromethane,washed with a saturated aqueous solution of sodium bicarbonate, brinethen the organic phase was dried (Na₂SO₄), filtered and evaporated.[(3R,4R,5S)-4-(Hydroxy)-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[ethyl-2-ene-Sar]³cyclosporin A (Intermediate 27) was obtained asa white solid after purification by SCX chromatography using a solventgradient of 100% methanol→0.14M ammonia in methanol.

ESMS MH⁺ 1287.2

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.23 (d, 1H, amide NH),7.72 (d, 1H, amide NH), 7.81 (d, 1H, amide NH).

Alternative compounds of Formula I may be produced according to SchemeVIII through the use of alternative heterocycles or amines at Step 3 inScheme VIII.

Exemplary compounds that may be used in place of morpholine at step 3 inscheme VIII to produce compounds of the present invention include butare not limited to 2 methyloctahydropyrrolo[3,4c]pyrrole, piperidine,N-methyl piperazine, homomorpholine, and pyrrolidine.

Preparation of Compound AB

A solution of[(3R,4R,5S)-4-(hydroxy)-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[ethyl-2-ene-Sar]³cyclosporin A (Intermediate 27) (0.187 g, 0.145mmol) in methanol (20 mL) was passed twice through a 10% palladium oncarbon cartridge in a H-cube system at 30° C. under 30 atm of hydrogen.The reaction mixture was concentrated and the residue purified by MPLCchromatography using a solvent gradient of 100% dichloromethane→95%dichloromethane/5% methanol containing 10% aqueous ammonia (0.88) togive[(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-ethyl-Sar]³cyclosporin A (Compound AB) as a white solid.

H-cube is a continuous flow reactor for carrying out hydrogenation underpressure. The hydrogenation is carried out using palladium on carboncatalyst at a hydrogen pressure of 30 atmospheres.

ESMS MH⁺ 1289.5

¹H NMR (CDCl₃, ppm) 7.19 (d, 1H, amide NH), 7.37 (d, 1H, amide NH), 7.80(d, 1H, amide NH), 7.98 (d, 1H, amide NH).

Scheme IX Procedure for Obtaining a Compound Having Formula I, WhereinR¹ is —SCH₃, R¹¹ is 0, n=0, m=0 and p=1, and wherein R⁹, R¹⁰, R¹¹, andthe N to which R⁹ and R¹⁹ are attached taken together form

wherein “*” Represents the Point of Attachment to R⁵. Scheme IX

Preparation of Intermediate 17 (Compound A)

[(3R,4R,5S)-4-(hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹cyclosporin A (Intermediate 17, Compound A) is prepared accordingto steps 2 and 3 of Scheme VIII using morpholine as the reactant in step3.

ESMS MH⁺ 1261.59

¹H NMR (CDCl₃, ppm) δ 7.17 (d, 1H, amide NH), 7.40 (d, 1H, amide NH),7.74 (d, 1H, amide NH), 7.91 (d, 1H, amide NH)

Compound LL (see table 2)[(3R,4R,5S)-4-(hydroxy-3-methyl-5-(methylamino)-1-N-methylpiperazinyl-hexanoicacid]¹cyclosporin A

was prepared in a similar manner by using N-methylpiperazine as thereactant in step 3.

ESMS MH⁺ 1274.64

¹H NMR (CDCl₃, ppm) δ 7.17 (d, 1H, amide NH), 7.39 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 7.92 (d, 1H, amide NH)

Compound NN (see table 3)[(3R,4R,5S)-4-(hydroxy-3-methyl-5-(methylamino)-1-N-methylpiperazinyl-hexanoicacid]¹cyclosporin A

was prepared in a similar manner by using ketopiperazine as the reactantin step 3.

ESMS MH⁺ 1274.71

¹H NMR (CDCl₃, ppm) δ 7.05 (d, 1H, amide NH), 8.05 (d, 1H, amide NH),8.35 (d, 1H, amide NH), 8.41 (d, 1H, amide NH).

Compound AN (see table 19)[(3R,4R,5S)-4-(hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹cyclosporin D

was prepared in a similar manner by using Cyclosporin D as startingmaterial and morpholine as the reactant in step 3.

ESMS MH⁺ 1275.78

¹H NMR (CDCl₃, ppm) 5.8 (s, 1H, (R)-Sar³-H), 8.0 (d, 1H, amide NH), 7.75(d, 1H, amide NH), 7.5 (d, 1H, amide NH), 7.15 (d, 1H, amide NH)

Preparation of Compound K

To a stirred solution of diisopropylamine (0.47 ml, 4.75 mmol) in THF(30 ml) at −78° C. under nitrogen was added dropwise n-butyl lithium(2.5M in hexane, 1.9 ml, 4.75 mmol) over 5 minutes. The reaction mixturewas stirred at −78° C. for 1 hour, then[(3R,4R,5S)-4-(hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹cyclosporin A (Intermediate 17) (0.6 g, 0.5 mmol) in THF (10 ml)was added dropwise over 5 minutes. The reaction mixture was stirred fora further 2 hours before methyl disulphide (0.42 ml, 4.75 mmol) wasadded and the reaction was allowed to warm to room temperature over 18h. After this time the reaction mixture was quenched with ammoniumhydroxide solution (100 ml) then extracted with ethyl acetate (2×50 ml).The combined organic phases were washed with saturated brine (100 ml),then dried (MgSO₄), filtered and evaporated. The crude orange solidobtained (0.4 g) was purified by passing through a 20 g silica columnusing 3% MeOH/dichloromethane as the eluent to obtain Compound K([(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-thiomethyl-Sar]³cyclosporin A) as an off white solid.

ESMS MH⁺ 1308.02

¹H NMR (CDCl₃, ppm) 5.86 (s, 1H, (R)-Sar³-H), 7.20 (m, 2H, 2× amide NH),7.81 (m, 2H, 2× amide NH).

Other Compounds of Formula I in which R¹ is —SC₂₋₄ alkyl may be preparedusing alternative disulphides in place of dimethyldisulphide (of MeSSMe)or by using other similar reagents such as MeSO₂SC₂₋₄ alkyl in place ofMeSSMe at Step 2 in Scheme IX.

Using the procedure shown in Scheme IX with isopropyl disulphide in Step2 Compound I was prepared

Compound I (see Table 10),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-thio-isopropyl-Sar]³cyclosporin A

ESMS MH⁺ 1335.41

¹H NMR (CDCl₃, ppm) δ 7.12 (d, 1H, amide NH), 7.21 (d, 1H, amide NH),7.81 (d, 1H, amide NH), 7.88 (d, 1H, amide NH).

Using the procedure shown in Scheme IX with Cyclosporin D as thestarting material instead of Cyclosporin A, Compound AM was prepared.

Compound AM (see Table 19),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-morpholino-hexanoicacid]¹[(S)-thio-methyl-Sar]³cyclosporin D

ESMS MH⁺ 1321.43

¹H NMR (CDCl₃, ppm) δ 7.9 (d, 1H, amide NH), 7.8 (d, 1H, amide NH), 7.3(d, 1H, amide NH), 7.2 (d, 1H, amide NH)

Preparation of Intermediate 18

Intermediate 18 is a structural variant of cyclosporin A having an —OCH₃at the position 3 α-carbon. Intermediate 18 can be used in place ofcyclosporin A according to any of the schemes shown herein to producecompounds of Formula I (e.g., amines and amides of Formula I) in whichR¹ is —OCH₃. Intermediate 18 can be prepared as described in Example 1((3-methoxycyclosporin) of US 2010/0167996. The chemical structure ofIntermediate 18 is shown below.

For example, Compound B can be prepared from Intermediate 18 as shownbelow

Compound B (see Table 4),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-methoxy-Sar]³cyclosporin A

ESMS MH⁺ 1291.9

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.19 (d, 1H, amide NH),7.79 (d, 1H, amide NH), 7.85 (d, 1H, amide NH).

Preparation of Intermediate 23

Intermediate 23 is a structural variant of cyclosporine A having a—CH₂OCH₃ at the position 3 α-carbon. Intermediate 23 can be used inplace of Cyclosporin A according to any of the schemes shown herein toproduce compounds of Formula I (e.g., amines and amides of Formula I) inwhich R¹ is —CH₂OCH₃. The synthetic scheme for preparing Intermediate 23is shown below.

[(D)-Serine]³cyclosporin A is prepared as described by D. Seebach et al.(1993) Helvetica Chimica Acta 73(4): 1564-1590. To[(D)-Serine]³cyclosporin A (350 mg, 0.28 mmol) dissolved indichloromethane (3 mL) was added benzyltriethylammonium chloride (65 mg,0.28 mmol) and aqueous KOH solution (31%, 5.1 mL). Iodomethane (18 μL,0.28 mmol) was added and the mixture stirred rapidly for 18 hours atroom temperature. The reaction mixture was diluted with water (5 mL) anddichloromethane (5 mL) and the layers separated. The aqueous layer wasfurther extracted with dichloromethane (3×10 mL) and the combinedorganic layers dried (MgSO₄) and evaporated in vacuo. The residue waspurified by silicagel chromatography using 6% methanol indichloromethane to provide [(R)-methoxymethylene-Sar]³cyclosporin A(Intermediate 23) as an off-white solid.

ESMS MH⁺ 1246.87

¹H NMR (CDCl₃, ppm) δ 7.17 (d, 1H, amide NH), 7.44 (d, 1H, amide NH),7.65 (d, 1H, amide NH), 8.03 (d, 1H, amide NH).

For example Compound D can be prepared from Intermediate 23 andsimilarly Compounds V, S and AD can be prepared from[(D)-Serine]³cyclosporin A as shown below.

Compound D (see Table 5),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-methoxymethylene-Sar]³cyclosporin A

ESMS MH⁺ 1305.8

¹H NMR (CDCl₃, ppm) δ 7.18 (d, 1H, amide NH), 7.37 (d, 1H, amide NH),7.77 (d, 1H, amide NH), 7.95 (d, 1H, amide NH).

Compound V (see Table 13),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-hydroxymethyl-Sar]³cyclosporin A

ESMS MH⁺ 1291.5

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.41 (d, 1H, amide NH),7.77 (d, 1H, amide NH), 7.99 (d, 1H, amide NH).

In a similar manner Compound EO and Compound EP can be prepared from[D-Serine]³cyclosporin A using 2,2,6,6-tetrafluoromorpholine or3,3-difluoropyrrolidine as reagents.

Compound EO (see table 13)(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2,2,6,6-tetrafluoro-morpholin-4-yl)-hexanoicacid]¹[D-Serine]³cyclosporin A

ES/MS: 1364.0 MH⁺

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.52 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 8.18 (d, 1H, amide NH).

Compound EP (see table 13)(3R,4R,5S)-1-(3,3-Difluoro-pyrrolidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[D-Serine]³cyclosporin A

ES/MS: 1311.3 MH⁺

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.42 (d, 1H, amide NH),7.77 (d, 1H, amide NH), 8.02 (d, 1H, amide NH).

Compound S (see Table 12),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-2 diethylamino ethyl oxymethyl-Sar]³cyclosporin A

ESMS MH⁺ 1390.7

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.48 (d, 1H, amide NH),7.76 (d, 1H, amide NH), 7.96 (d, 1H, amide NH).

Compound AD (see Table 14),[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-homomorpholino-hexanoicacid]¹[(R)-2 diethylamino ethyl oxymethyl-Sar]³cyclosporin A

ESMS MH⁺ 1404.7

¹H NMR (CDCl₃, ppm) δ 7.20 (d, 1H, amide NH), 7.39 (d, 1H, amide NH),7.78 (d, 1H, amide NH), 7.98 (d, 1H, amide NH).

Scheme X Procedure for Obtaining a Compound Having Formula I, Wherein R¹is —CH₃, R² is —CH₃, R³ is —CH₂CH(CH₃)₂, R⁴ is —CH₃, R⁵ is —CH₂CH₂CH₂—,R⁶ is —CH₂CH₃, R⁸ is CH₂, n=2, m=0, and p=0

Schemes X and Xa describe processes for the synthesis of amines ofFormula I.

Scheme X describes the acylation of Intermediate 1 (Step 1) andbromination of Intermediate 29 by N-bromosuccinimideazobisisobutyrylnitrile in carbon tetrachloride (Step 2) carried out asdescribed by M K Eberle et al. J. Org. Chem. 1992, 57, 2689-2691. Steps3 and 4 are carried out as described in Steps 2 and 3 in Scheme II.Intermediate 1 is obtained as described in Scheme I. Step 5 is carriedout by stirring Intermediate 32 in a solution of potassium carbonate ina mixture of methanol and water as described for Example 8 in US2003/0212249.

Scheme Xa

Scheme Xa describes the use of Wittig chemistry on Intermediate 4 (Step1).

Intermediate 4 is obtained as described in Scheme I. Deprotection ofIntermediate 35 and hydrogenation of Intermediate 36 are carried out asdescribed in Steps 5 and 3, respectively, in Scheme I. Exemplarysyntheses according to Scheme Xa are described below for the preparationof Compounds KF and KG.

Scheme Xa (Example 1) Preparation of Compound KF

Step 1 Preparation of Intermediate 35

To an ice cooled suspension of(2-dimethylaminoethyl)triphenylphosphonium bromide (435 mg, 1.05 mmol)in dry tetrahydrofuran (6 mL) was added a solution of 2.5M n-butyllithium in hexanes. The deep orange suspension was stirred undernitrogen for 50 minutes before the addition of a solution of[(3R,4R,5S)-4-(t-butyldimethylsilanyloxy)-3-methyl-5-(methylamino)-1-oxo-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 4) (198 mg, 0.15mmol) in dry tetrahydrofuran (2 mL). The resulting yellow suspension wasstirred at room temperature for 67 hours then concentrated in vacuo. Theresidue was partitioned between diethyl ether and water then the organicphase was dried over sodium sulphate and concentrated in vacuo.[(2E,5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-1-(dimethylamino)-5-methyl-7-(methylamino)-oct-2-enoic acid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate35) was obtained as a white solid after purification by SCXchromatography using a solvent gradient of 100% methanol→0.35M ammoniain methanol and used as such in the next step.

¹H NMR (CDCl₃, ppm) δ 7.20 (d, 1H, amide NH), 7.93 (d, 1H, amide NH),8.07 (d, 1H, amide NH), 8.57 (d, 1H, amide NH).

Step 2 Preparation of Intermediate 36

To a stirred solution of[(2E,5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-1-(dimethylamino)-5-methyl-7-(methylamino)-oct-2-enoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 35) (0.166 g, 0.12mmol) in dry tetrahydrofuran (5 ml) was added tetrabutylammoniumfluoride TBAF (1.0M solution in tetrahydrofuran, 0.6 ml, 0.6 mmol) andthe yellow solution stirred at room temperature for 18 hours. Thesolvent was evaporated, the residue obtained dissolved in ethyl acetateand washed with H₂O. The organic phase was dried (Na₂SO₄), filtered andevaporated. The residue was purified by MPLC chromatography using asolvent gradient of 100% dichloromethane→95% dichloromethane/5% methanolcontaining 10% aqueous ammonia (0.88) to give[(2E,5R,6R,7S)-1-(dimethylamino)-6-hydroxy-5-methyl-7-(methylamino)-oct-2-enoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 36) as a white solid.

ESMS MH⁺ 1259.5

¹H NMR (CDCl₃, ppm) 7.21 (d, 1H, amide NH), 7.51 (d, 1H, amide NH), 7.74(d, 1H, amide NH), 8.08 (d, 1H, amide NH).

Step 3 Preparation of Compound KF

[(2E,5R,6R,7S)-1-(Dimethylamino)-6-hydroxy-5-methyl-7-(methylamino)-oct-2-enoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 36) (50 mg, 0.04mmol) was dissolved in ethanol (10 mL), treated with 10% palladium oncarbon (25 mg) then placed under a 1 atm atmosphere of hydrogen for 22hours. The reaction mixture was filtered through celite thenconcentrated in vacuo.[(5R,6R,7S)-1-(dimethylamino)-6-hydroxy-5-methyl-7-(methylamino)-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound KF) was obtained as awhite solid after purification by SCX chromatography using a solventgradient of 100% methanol→0.18M ammonia in methanol.

ESMS MH⁺ 1261.2

¹H NMR (CDCl₃, ppm) δ 7.13 (d, 1H, amide NH), 7.49 (d, 1H, amide NH),7.68 (d, 1H, amide NH), 7.93 (d, 1H, amide NH).

Scheme Xa (Example 2) Preparation of Compound KG

Step 1 Preparation of Intermediate 37

Using the procedure described above for the preparation of Intermediate35 with (2-(N-morpholino) ethyl) triphenylphosphonium bromide (G. V. Raoet al. Tetrahedron Lett. 49 (2008) 824) as starting material,[(2E,5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-1-(N-morpholino)-oct-2-enoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 37) was obtained as awhite solid after purification by SCX chromatography using a solventgradient of 100% methanol→0.14M ammonia in methanol and used as such inthe next step.

ESMS MH⁺ 1415.7

Step 2 Preparation of Intermediate 38

Using the procedure described above for the preparation of Intermediate36 with[(2E,5R,6R,7S)-6-(t-butyldimethylsilanyloxy)-5-methyl-7-(methylamino)-1-(N-morpholino)-oct-2-enoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 37) as startingmaterial,[(2E,5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(N-morpholino)-oct-2-enoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 38) was obtained as awhite solid after purification by SCX chromatography using a solventgradient of 100% methanol→0.35M ammonia in methanol.

ESMS MH⁺ 1301.3

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.52 (d, 1H, amide NH),7.72 (d, 1H, amide NH), 8.06 (d, 1H, amide NH).

Step 3 Preparation of Compound KG

Using the procedure described above for the preparation of Compound KF(Step 3) with[(2E,5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(N-morpholino)-oct-2-enoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Intermediate 38) as startingmaterial,[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(N-morpholino)-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound KG) was obtained as awhite solid after purification by SCX chromatography using a solventgradient of 100% methanol→0.35M ammonia in methanol.

ESMS MH⁺ 1303.4

¹H NMR (CDCl₃, ppm) δ 7.13 (d, 1H, amide NH), 7.50 (d, 1H, amide NH),7.67 (d, 1H, amide NH), 7.91 (d, 1H, amide NH).

Scheme XI Procedure for Obtaining a Compound Having Formula I, WhereinR¹ is —R¹³R¹⁴, R¹¹ is O, R¹¹ is CH₂S, R¹⁴ is —CH₂CH₂N(CH₂CH₃)₂, n=0, m=0and p=1, and wherein R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areAttached Taken Together Form a Heterocycle

Intermediate 3 is prepared as described in Scheme I. Intermediate 33 wasprepared as described in Step 4 in Scheme I. Intermediate 34 wasprepared as described in Step 5 in Scheme I.

Compound AC[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-2-diethylaminoethylthiomethyl-Sar]³cyclosporinA was preparedin the following manner.

To a solution of[(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[methylene-Sar]³cyclosporinA (0.130 g, 0.1 mmol) (Intermediate 34)in acetonitrile (10 ml) was added cesium carbonate (0.332 g, 1 mmol) andthe white suspension was bubbled through with nitrogen for one hourbefore adding diethylaminoethylthiol hydrochloride salt (0.087 g, 0.51mmol). The reaction mixture was bubbled through with nitrogen overnightduring which time the solvent had evaporated. Fresh acetonitrile (15 ml)and additional amounts of cesium carbonate (0.332 g, 1 mmol) were added,the resulting suspension was bubbled through with nitrogen for one hourbefore adding additional amounts of diethylaminoethylthiol hydrochloridesalt (0.087 g, 0.51 mmol). The reaction mixture was bubbled through withnitrogen overnight during which time the solvent had evaporated. Freshacetonitrile was added and the suspension filtered through a pad ofsodium sulphate then concentrated to give 170 mg of a colourless oil.Purification by MPLC chromatography using a solvent gradient of 100%dichloromethane→95% dichloromethane/5% methanol containing 10% aqueousammonia (0.88) followed by trituration with hexane in ultrasonic bathprovided[(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-2-diethylamino ethyl thiomethyl-Sar]³cyclosporin A (CompoundAC).

ESMS MH⁺ 1406.5

¹H NMR (CDCl₃, ppm) δ 7.18 (1H, d, amide NH), 7.30 (1H, d, amide NH),7.79 (1H, d, amide NH), 8.02 (1H, d, amide NH).

Other thiols such as morpholinoethylthiol, diethylaminopropylthiol,ethanethiol and morpholinopropylthiol may be reacted with Intermediate34 in a similar manner to give the corresponding compounds of Formula I.

Preparation of[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DA)

ESMS MH⁺ 1310.8

¹H NMR (CDCl₃, ppm) δ 7.20 (d, 2H, amide NH and pyridine CH), 7.45 (d,1H, amide NH), 7.52 (m, 1H, pyridine CH), 7.69 (dd, 1H, pyridine CH),7.80 (d, 1H, amide NH), 7.95 (d, 1H, amide NH), 8.53 (d, 1H, pyridineCH).

Compound DA is prepared from intermediate 4 according to Scheme I andthe steps shown below. Compounds DB, DC, DD, DE, DF, DG, DH, DI, DJ, DK,DL, DM, DN, DO, DP, DQ, and DR are prepared in a similar manneraccording to Scheme I. The structures for each of these compounds alongwith their corresponding proton NMR and electron spray mass spectroscopydata are given below.

[(3R,4R,5S)-1-(Bis{pyridin-2-ylmethyl}amino)-4-hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DB)

ESMS MH⁺ 1387.8

¹H NMR (CDCl₃, ppm) δ 7.13 (m, 3H, amide 1NH and pyridine 2CH), 7.52 (m,3H, amide 1NH and pyridine 2CH), 7.71 (m, 3H, amide 1NH and pyridine2CH), 8.02 (d, 1H, amide NH), 8.53 (d, 2H, pyridine CH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(methyl-phenyl-amino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DC)

ESMS MH⁺ 1295.8

¹H NMR (CDCl₃, ppm) δ 6.66 (m, 3H, phenyl CH), 7.11 (d, 1H, amide NH),7.19 (dd, 2H, phenyl CH), 7.50 (d, 1H, amide NH), 7.70 (d, 1H, amideNH), 7.97 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(methyl-pyridin-2-yl-amino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DD)

ESMS MH⁺ 1296.7

¹H NMR (CDCl₃, ppm) δ 6.49 (dd, 2H, pyridine CH), 7.17 (d, 1H, amideNH), 7.42 (dd, 1H, pyridine NH), 7.48 (d, 1H, amide NH), 7.80 (d, 1H,amide NH), 8.02 (d, 1H, amide NH), 8.1 (d, 1H, pyridine CH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-sulfamoyl-ethyl)-methyl-amino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DE)

ESMS MH⁺ 1326.7

¹H NMR (CDCl₃, ppm) δ 7.12 (d, 1H, amide NH), 7.53 (d, 1H, amide NH),7.71 (d, 1H, amide NH), 8.06 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-3-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DF)

ESMS MH⁺ 1310.5

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.25 (m, 1H, pyridine CH),7.45 (d, 1H, amide NH), 7.70 (bd, 1H, pyridine CH), 7.74 (d, 1H, amideNH), 7.91 (d, 1H, amide NH), 8.48 (bs, 2H, pyridine CH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyrimidin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DG)

ESMS MH⁺ 1311.7

¹H NMR (CDCl₃, ppm) δ 7.18 (m, 2H, amide NH and pyrimidine CH), 7.42 (d,1H, amide NH), 7.76 (d, 1H, amide NH), 7.93 (d, 1H, amide NH), 8.72 (d,2H, pyrimidine CH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyrazin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DH)

ESMS MH⁺ 1311.8

¹H NMR (CDCl₃, ppm) δ 7.14 (d, 1H, amide NH), 7.48 (d, 1H, amide NH),7.74 (d, 1H, amide NH), 7.92 (d, 1H, amide NH), 8.48 (d, 2H, pyrazineCH), 8.7 (s, 1H, pyrazine CH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3-methyl-3H-imidazol-4-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DI)

ESMS MH⁺ 1313.6

¹H NMR (CDCl₃, ppm) δ 6.86 (s, 1H, imidazole CH), 7.12 (d, 1H, amideNH), 7.39 (s, 1H, imidazole CH), 7.49 (d, 1H, amide NH), 7.72 (d, 1H,amide NH), 7.93 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({2-methyl-2H-pyrazol-3-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DJ)

ESMS MH⁺ 1313.6

¹H NMR (CDCl₃, ppm) δ 6.10 (d, 1H, pyrazole CH), 7.12 (d, 1H, amide NH),7.36 (d, 1H, pyrazole CH), 7.48 (d, 1H, amide NH), 7.72 (d, 1H, amideNH), 7.92 (d, 1H, amide NH).

[(3R,4R,5S)-1-({2-Cyano-propyl}-methyl-amino)-4-hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DK)

ESMS MH⁺ 1286.8

¹H NMR (CDCl₃, ppm) δ 7.12 (d, 1H, amide NH), 7.49 (d, 1H, amide NH),7.70 (d, 1H, amide NH), 7.95 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-4-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DL)

ESMS MH⁺ 1310.7

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.25 (d, 2H, pyridine CH),7.47 (d, 1H, amide NH), 7.75 (d, 1H, amide NH), 7.92 (d, 1H, amide NH),8.52 (d, 2H, pyridine CH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({1-methyl-1H-pyrazol-4-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DM)

ESMS MH⁺ 1313.6

¹H NMR (CDCl₃, ppm) δ 7.21 (d, 1H, amide NH), 7.31 (s, 1H, pyrazole CH),7.35 (s, 1H, pyrazole CH), 7.43 (d, 1H, amide NH), 7.84 (d, 1H, amideNH), 7.92 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3,3,3-trifluoropropyl}-methyl-amino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DN)

ESMS MH⁺ 1315.8

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.45 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 7.91 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({1-methyl-3-trifluoromethyl-2H-pyrazol-5-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DO)

ESMS MH⁺ 1381.7

¹H NMR (CDCl₃, ppm) δ 6.38 (s, 1H, pyrazole CH), 7.11 (d, 1H, amide NH),7.51 (d, 1H, amide NH), 7.70 (d, 1H, amide NH), 7.96 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({5-fluoro-pyridin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DP)

ESMS MH⁺ 1328.5

¹H NMR (CDCl₃, ppm) δ 7.17 (d, 1H, amide NH), 7.48 (m, 3H, amide 1NH andpyridine 2CH), 7.73 (d, 1H, amide NH), 7.94 (d, 1H, amide NH), 8.38 (d,1H, pyridine CH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({5-chloro-pyridin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DQ)

ESMS MH⁺ 1344.6

¹H NMR (CDCl₃, ppm) δ 7.16 (d, 1H, amide NH), 7.48 (m, 2H, amide NH andpyridine CH), 7.68 (m, 1H, pyridine CH), 7.73 (d, 1H, amide NH), 7.93(d, 1H, amide NH), 8.48 (d, 1H, pyridine CH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3-trifluoromethyl-pyridin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DR)

ESMS MH⁺ 1378.5

¹H NMR (CDCl₃, ppm) δ 7.14 (d, 1H, amide NH), 7.47 (d, 1H, amide NH),7.54 (d, 1H, pyridine CH), 7.73 (m, 2H, amide NH and pyridine CH), 7.90(m, 2H, amide NH and pyridine CH).

Compounds DS, DT, DU, DV, DW, DX, DZ, EA, EB, EC, ED, and EE areprepared by reaction of intermediate 4 with the appropriate cyclic amineaccording to Scheme I.The structures and corresponding proton NMR and mass spectroscopy datafor each compound are shown below.

[(3R,4R,5S)-1-(3,3-Dimethyl-morpholin-4-yl)-4-hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DS)

ESMS MH⁺ 1303.77

¹H NMR (CDCl₃, ppm) δ 7.13 (d, 1H, amide NH), 7.41 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 7.82 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-methylamino-((R)-3-methyl-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DT)

ESMS MH⁺ 1289.81

¹H NMR (CDCl₃, ppm) δ 7.14 (d, 1H, amide NH), 7.45 (d, 1H, amide NH),7.72 (d, 1H, amide NH), 7.91 (d, 1H, amide NH).

[(3R,4R,5S)-1-(5,6-Dihydro-8H-[1,2,4]triazolo[4,3-a]pyrazin-7-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DU)

ESMS MH⁺ 1312.89

¹H NMR (CDCl₃, ppm) δ 7.22 (d, 1H, amide NH), 7.51 (d, 1H, amide NH),7.81 (d, 1H, amide NH), 8.05 (d, 1H, amide NH), 8.10 (s, 1H, aromaticCH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(8-oxa-3-aza-bicyclo[3.2.1]oct-3-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DV)

ESMS MH⁺ 1301.85

¹H NMR (CDCl₃, ppm) δ 7.18 (d, 1H, amide NH), 7.51 (d, 1H, amide NH),7.77 (d, 1H, amide NH), 7.94 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-((S)-3-methyl-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DW)

ESMS MH⁺ 1289.80

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.40 (d, 1H, amide NH),7.75 (d, 1H, amide NH), 7.83 (d, 1H, amide NH).

[(3R,4R,5S)-1-(2,3-Dihydro-benzo[1,4]oxazin-4-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DX)

ESMS MH⁺ 1323.73

¹H NMR (CDCl₃, ppm) δ 7.12 (d, 1H, amide NH), 7.51 (d, 1H, amide NH),7.71 (d, 1H, amide NH), 7.99 (d, 1H, amide NH).

[(3R,4R,5S)-1-(1,4-Dioxa-8-aza-spiro[4.5]dec-8-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DY)

ESMS MH⁺ 1332.09

¹H NMR (CDCl₃, ppm) δ 7.20 (d, 1H, amide NH), 7.37 (d, 1H, amide NH),7.79 (d, 1H, amide NH), 7.87 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-phenyl-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DZ)

ESMS MH⁺ 1352.49

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.31-7.38 (m, 5H, AromaticCH), 7.39 (d, 1H, amide NH), 7.47 (d, 1H, amide NH), 7.77 (d, 1H, amideNH), 7.78 (d, 1H, amide NH), 7.93 (d, 1H, amide NH), 8.01 (d, 1H, amideNH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(piperidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EA)

ESMS MH⁺ 1273.69

¹H NMR (CDCl₃, ppm) δ 7.23 (d, 1H, amide NH), 7.37 (d, 1H, amide NH),7.84 (d, 1H, amide NH), 7.90 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(pyrrolidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EF)

ESMS MH⁺ 1259.78

¹H NMR (CDCl₃, ppm) δ 7.21 (d, 1H, amide NH), 7.33 (d, 1H, amide NH),7.81 (d, 1H, amide NH), 7.83 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-trifluoromethyl-piperidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EB)

ESMS MH⁺ 1341.16

¹H NMR (CDCl₃, ppm) 2 isomers δ 7.13 (d, 1H, amide NH), 7.18 (d, 1H,amide NH), 7.45 (d, 1H, amide NH), 7.47 (d, 1H, amide NH), 7.70 (d, 1H,amide NH), 7.72 (d, 1H, amide NH), 7.95 (d, 1H, amide NH), 7.97 (d, 1H,amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(3-trifluoromethyl-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EC)

ESMS MH⁺ 1344.91

¹H NMR (CDCl₃, ppm) 2 isomers δ 7.11 (d, 1H, amide NH), 7.12 (d, 1H,amide NH), 7.49 (d, 1H, amide NH), 7.51 (d, 1H, amide NH), 7.67 (d, 1H,amide NH), 7.68 (d, 1H, amide NH), 7.97 (d, 1H, amide NH), 7.99 (d, 1H,amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-([1,2]oxazinan-2-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound ED)

ESMS MH⁺ 1276.43

¹H NMR (CDCl₃, ppm) δ 7.22 (d, 1H, amide NH), 7.40 (d, 1H, amide NH),7.79 (d, 1H, amide NH), 8.05 (d, 1H, amide NH).

[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-trifluoromethyl-5,6-dihydro-8H-imidazo[1,2-a]pyrazin-7-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EE)

ESMS MH⁺ 1379.86

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, aromatic CH), 7.17 (d, 1H, amide NH),7.51 (d, 1H, amide NH), 7.75 (d, 1H, amide NH), 7.98 (d, 1H, amide NH).

Compounds EG, EH, and EI are prepared according to Scheme I. Structuresfor each are shown below.

[[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(4-methyl-[1,4]diazepan-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EG)

ESMS MH⁺ 1302.5

¹H NMR (CDCl₃, ppm) δ 7.18 (d, 1H, amide NH), 7.46 (d, 1H, amide NH),7.74 (d, 1H, amide NH), 7.96 (d, 1H, amide NH).

[[(3R,4R,5S)-4-Hydroxy-1-(3-methoxy-azetidin-1-yl)-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EH)

ESMS MH⁺ 1275.9

¹H NMR (CDCl₃, ppm) δ 7.19 (d, 1H, amide NH), 7.36 (d, 1H, amide NH),7.78 (d, 1H, amide NH), 7.88 (d, 1H, amide NH).

Compound[(4R,5R,6S)-5-Hydroxy-4-methyl-6-(methylamino)-1-(morpholin-4-yl)-heptanoicacid]¹cyclosporin A (Compound EI)

ESMS MH⁺ 1275.46

¹H NMR (CDCl₃, ppm) δ 7.15 (d, 1H, amide NH), 7.49 (d, 1H, amide NH),7.69 (d, 1H, amide NH), 8.03 (d, 1H, amide NH).

Compound EI is prepared from an intermediate similar to intermediate 15by using Scheme Xa in a manner similar to that used to prepare CompoundKG, as shown below.

Intermediate 101 is prepared from CsA in a manner similar to that usedto prepare intermediate 15 in Scheme VI, as shown by the scheme shownbelow.Intermediate 104 is prepared in a similar manner to intermediate 2 inScheme I.Intermediate 105 is prepared in a similar manner to intermediate 3 inScheme I.Intermediate 106 is prepared in a similar manner to intermediate 14 inScheme VI.

[(3R,4R,5S)-4-Hydroxy-3-methyl-1-(morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³[Ethyl-Val]⁴cyclosporin A (Compound EJ)

ESMS MH⁺ 1275.77

¹H NMR (CDCl₃, ppm) δ 7.06 (d, 1H, amide NH), 7.17 (d, 1H, amide NH),7.76 (d, 1H, amide NH), 7.79 (d, 1H, amide NH).

Biological and Physical Properties

Example Compounds of Formula I are listed and described in Tables 1-27,below. Compounds of the present invention include those listed anddescribed in Tables 1-27, below, and their pharmaceutically acceptablesalts.

Data Showing Cyclophilin A (Cyp A) Inhibitory Activity,immunosuppressive Potential, and aqueous solubility for select Compoundshaving Formula I are described in Tables 28-35. General procedures andassays used to obtain the data are given below.

The data shows that many compounds having Formula I, are potentinhibitors of cyclophilin A (Ki<10 nM), as measured by the protease-freePPIase assay. Many compounds having Formula I are alsonon-immunosuppressive, as measured by the MLR and CaN assays; and aremore water soluble than unmodified Cyclosporin A, as measured by theWater Solubility Assay. Surprisingly, however, a sub-set of compoundshaving Formula I show more immunosuppressive potential than others.Examples include Compounds EK, EL, EM, EN, EQ, ER, EO, and EP.

General Procedures and Assays *Protease-Free PPlase Assay

The protease-free PPIase assay measures the rate of cis to transconversion of a peptide substrate catalyzed by the enzyme cyclophilin A.Addition of a cyclophilin A inhibitor (e.g., a test compound) slows thecatalyzed rate and a K_(i) value is obtained. A K_(i) value of less than10 nM demonstrates that the test compound is a potent inhibitor ofcyclophilin A.

Materials

Assay Buffer:

35 mM HEPES pH 7.8, filtered through a 0.2 μm filter. 50 μM DTT wasadded prior to use each day and then the buffer was stored on ice.

Enzyme:

Human recombinant cyclophilin A (Cyp A) (Sigma C3805) enzyme was dilutedto 1 μM with enzyme dilution buffer (20 mM HEPES pH 7.8, 40% glycerol,50 μM DTT and 1 μM BSA) and stored at −20° C.

Substrate:

Succinimide-Ala-Ala-Pro-Phe-p-nitroanailide (SUC-AAPF-pNA) (from BachemAG, L-1400), 20 mg/ml prepared in 0.5 M LiCl in trifluoroethanol.

Method

All readings were taken with an Agilent 8453 Spectrophotometer whichconsists of a cuvette holder, stirrer and chiller to maintain a stirredcuvette temperature of 10.0±0.1° C. The temperature is monitored by theuse of temperature probe. To prevent UV degradation of test compounds,the light below 290 nm was blocked using a glass slide in the lightpath. 1.5 ml of assay buffer was put into a 3 ml quartz cuvette andcooled to 10.0±0.1° C. while stirring (vigorous but not so fast as toproduce cavitation). The inhibitor was diluted in 100% DMSO, and thenadded to the assay to a maximum final concentration of 0.5% DMSO in theassay. A blank spectrum was obtained, then 3 μL of enzyme was added (2nM final concentration) and then 3 μL substrate (60 μM finalconcentration) added. The absorbance was measured at 330 nm for 300 s or500 s for blank runs (NOTE: the substrate must be added in one quickinjection and the measurements started immediately to minimize mixingerrors).

A first order rate equation was fitted to the absorbance data, for eachconcentration of inhibitor, to obtain the rate constant (the first 10 to15 seconds were excluded as mixing causes errors in this portion ofcurve). The catalytic rate was calculated from the enzymatic rateconstant minus the background rate constant. An exponential curve wasgenerated using the catalytic rate constants versus the inhibitorconcentration to obtain the K_(i) value for the inhibitor. The K_(i)value is indicative of the binding affinity between the test compoundand cyclophilin A.

**Calcineurin Phosphatase (CaN) Assay

The calcineurin phosphatase assay is a means for estimating theimmunosuppressive potential of a test compound. Calcineurin is aserine-threonine protein phosphatase that on activation dephosphorylatesmembers of the nuclear factor of activated T cells (NFAT), which areimportant in T lymphocyte activation. Cyclosporin A (CsA) bound tocyclophilin A (Cyp A) inhibits calcineurin activity, thus resulting inimmunosuppressive effects. Although CsA only inhibits calcineurin whenbound to Cyp A, some Cyclosporin A (CsA) analogs will also bindcalcineurin in the absence of Cyp A. Alternatively, some CsA analogsbind cyclophilin A but do not inhibit calcineurin activity.

To investigate the immunosuppressive potential of exemplary compounds ofFormula I, which are cyclosporin analogs, their ability to inhibitcalcineurin activity was measured in the presence and absence of Cyp A.

The CaN assay kit used is based on a colorimetric assay for measuringcalcineurin phosphatase activity, and it is commercially available (EnzoLife Sciences and Calbiochem). Calmodulin is also required forcalcineurin activity and RII phosphopeptide is used as an efficientpeptide substrate for calcineurin. We have modified the method to enablemeasurement of Cyp A-dependent and Cyp A-independent inhibition ofcalcineurin through the addition of Cyp A in a 1:1 complex with theinhibitor. The detection of free phosphate released is based on theclassic Malachite green assay.

Materials:

Enzo Life Sciences CaN Assay Kit: BML-AK804

2X Assay Buffer:

100 mM Tris, pH7.5, 200 mM NaCl, 12 mM MgCl₂, 1 mM DTT, 0.05% NP-40, 1mM CaCl₂

Malachite Green:

BIOMOL Green™ reagent

Calmodulin (Human, Recombinant):

was thawed on ice, diluted 1:50 with 2× assay buffer, and then stored onice.

Calcineurin:

was thawed quickly, stored on ice immediately, diluted 1:12.5 with 1×assay buffer, and then stored on ice.

R-II Substrate:

915 μL ultrapure water (UPW) was added to the 1.5 mg vial substrate togive a final concentration of 0.75 mM.

Inhibitors:

2.5 mM inhibitor in 100% DMSO.

Cyp A:

recombinant human CypA (Sigma C3805), 1 mg/ml; Recombinant 6-his taggedCypA prepared by the Univ. of Edinburgh was also used. Comparison of theresults showed that both enzymes gave identical results.

Method

Inhibitor Dilutions:

inhibitor compounds were diluted in UPW in polypropylene low-binding 96well plates at 5× the final assay concentration. For samples ‘withoutCyp A’, a 4-point dilution series of the inhibitor was prepared induplicate to obtain a final assay concentration of 10, 1, 0.1 and 0.01μM. For samples ‘with Cyp A’, a 7-point dilution series was prepared toobtain a 1:1 complex of the inhibitor with CypA; the inhibitor and Cyp Afinal assay concentrations of 10, 3.33, 1.11, 0.37, 0.12, 0.04, 0.014 μMwere prepared. Cs A inhibitor controls were also prepared to obtain afinal concentration of 10 μM Cs A with and without 10 μM Cyp A.

Assay Setup:

using the half area 96 well plates supplied with the kit, 10 μl UPW wasadded to duplicate wells to provide the non-inhibited control. 10 μl ofthe inhibitor or the inhibitor/Cyp A complex was added to theappropriate sample wells. 25 μl of the 2× assay buffer with CaM wasadded to all wells, then 5 μl of CaN was added to all wells (40 U perwell final concentration) except duplicate ‘no calcineurin blank’ wellsto which 50 μL, 1× assay buffer was added. The assay plate was placed inan oven at 30° C. for 15 minutes to equilibrate to the reactiontemperature. The reaction was started by the addition of 10 μlRII-peptide (0.15 mM final concentration). The reaction was allowed toproceed at 30° C. for a time period in which the reaction is linear forabout 60 minutes. The reaction was then terminated by adding 100 μl ofthe Malachite Green reagent. The color was allowed to develop for 15-30minutes at room temperature before the absorbance at 620 nm was measuredusing a plate reader (Molecular Devices—SpectraMax M5). The data wereanalyzed by subtracting ‘no Calcineurin blank’ from all the absorbancereadings and plotting the background corrected absorbances against Log₁₀inhibitor concentration. A sigmoidal-dose response curve was fitted tothe data using GraphPad Prism Software.

Cyclosporin A is a potent inhibitor of calcineurin activity andtherefore a potent immunosuppressive. It exerts its immunosuppressiveactivity by binding to cyclophilin A to form a complex, which then bindsto calcineurin and thereby inhibits calcineurin activity. As shown inthe tables, Cyclosporin A has a IC₅₀ value of 210 nM in thecalcineurin/cyclophilin A assay. Thus, compounds with values higher than210 nM in this assay will be predictably less immunosuppressive thancyclosporin A. As can be seen from the tables, many compounds of FormulaI produce much higher values than 210 nM in this assay and so would beexpected to be much less immunosuppressive than cyclosporin A. However,a sub-set of compounds of Formula I show immunosuppressive activity asmeasured by the Calcineurin phosphatase assay (IC₅₀ values between 225nM and 4 μM)

***Mixed Lymphocyte Reaction (“MLR”) Assay

The MLR assay is widely used in the field of immunology to measure Tcell proliferation, and therefore is another means of estimating theimmunosuppressive potential of test compounds. In the MLR assay,splenocytes isolated from two different strains of mice, termedStimulator (e.g. BALB/c mice) and Responder (e.g. C57BL/6 mice), aremixed in cell culture, in turn eliciting an alloimmune response(immunity against antigens between individuals of the same species).Alloimmunity results in robust proliferation of T cells contained withinthe splenocyte cell population from both strains of mice. To ensure thatT cell proliferation is restricted to only the Responder population(C57BL/6), the Stimulator cells (BALB/c) are first inactivated viax-irradiation before co-culture with Responder cells in the absence orpresence of different concentrations of test compound. If the testcompound present in the culture medium is immunosuppressive theproliferation of the responder cells is reduced. Total proliferation isquantified by the cellular uptake of [³H]-thymidine, which occurs duringcell division. Therefore, compounds that are less immunosuppressive thanCsA will require a higher concentration to reduce T cell proliferation;and compounds that are not immunosuppressive will not affect T cellproliferation even at the highest concentrations tested.

Female C57BL/6 and BALB/c mice, 6-8 weeks of age, were obtained from theFrederick Cancer Research and Development Center of the National CancerInstitute (Frederick, Md.). Spleens were harvested aseptically from allmice and single cell suspensions were prepared by disaggregating thecells with frosted glass slides, allowing the debris to settle, andwashing the cells twice with complete medium. Complete medium consistsof RPMI 1640 medium containing 25 mM HEPES buffer (HyClone, Logan, Utah)supplemented with 10% heat-inactivated fetal bovine serum (FBS; AtlantaBiologicals, Lawrenceville, Ga.), 100 μg/mL streptomycin, 100 U/mLpenicillin G, 0.25 μg/mL amphotericin B (HyClone), 2 mM L-glutaminedipeptide (HyClone), and 2×10⁻⁵ M 2-mercaptoethanol (Sigma). Cells werewashed twice and resuspended in complete medium. Cell counts wereperformed using a Beckman Coulter Z-1 particle counter (Fullerton,Calif.). Cell viability was determined by propidium iodide (PI) stainingusing an Accuri C6 flow cytometer (Ann Arbor, Mich.).

Spleen cells from C57BL/6 (H-2^(b)) and BALB/c (H-2^(d)) were used asresponder (R) and stimulator (S) cells, respectively. Cells were platedin triplicate in 96-well flat microtiter plates (Costar, Cambridge,Mass.) such that each well contained 2×10⁵ R and 8×10⁵ S cells. Cultureswere incubated in the absence or presence of various concentrations ofCsA, test compounds (e.g., a compound of Formula I), or medium at 37° C.in humidified 5% CO₂ for five days, pulsed with ³H-thymidine (³H-TdR)for the final 16 hours of incubation, and harvested using a Brandel96-well cell harvester (Gaithersburg, Md.). Proliferation was measuredby counting the radioactivity on filter mats in a Wallac 1450 MicrobetaTriLux scintillation counter (Turku, Finland). Controls to demonstrateeffective inactivation by the x-irradiation were performed by incubatingthe S cells with 5 μg/mL of PHA at 2×10⁵ cells/well. These controlcultures were incubated for 3 days under the same conditions as thosedescribed for the MLR; lymphoproliferation was determined in the samemanner as described above.

****Water Solubility Assay (Measured in pH 7.8 Buffer)

The aqueous solubility of a compound of Formula I in buffer (pH 7.8) wasmeasured by recording the onset of precipitation of the compound as afunction of increasing concentration. The onset of precipitation, if itoccurred, was detected by an increase in absorbance at 650 nm.

Materials

Assay Buffer:

35 mM HEPES pH 7.8

Stock solutions of Control and Test Compounds: 10 mM in 100% DMSO

Method

10 mM stock solutions of control and test compounds were prepared in100% DMSO. A series of dilutions were prepared from the stock in DMSO sothat the final concentrations in the assay were 0, 3.33, 10, 25, 50, 75and 100 μM and DMSO was limited to 1%.

Assay buffer (247.5 μl) was placed into flat bottomed transparent96-well plate. For blank samples DMSO (2.5 μl) was added. For test andcontrol samples 2.5 μl of the appropriate DMSO dilution stocks wereadded to the appropriate well. All test and control compounds wereperformed in triplicate.

The plates were sealed with adhesive plate seal and shaken at 250 rpm at25° C. for 18 h on a plate shaker.

After incubation the plate seals were taken off and any bubbles observedin wells removed. The plates were read on a SpectraMaxM5 with a 5 spre-shake at 650 nm.

Data files were transferred to the appropriate worksheet and thesolubility range of the compounds was calculated from the data.

The values shown in the tables indicate the concentration in μM(micromolar) at which the compound remains in solution.

TABLE 1 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q F —CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — A —HR¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 2 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q L —CH₃ R¹¹ is NR¹²; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ CH₃ 0 0 1 — LL —HR¹¹ is NR¹²; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachment to R⁵ CH₃ 0 0 1 —

TABLE 3 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q N —CH₃ R¹¹ is NR¹²; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ H 0 0 1 — NN —HR¹¹ is NR¹²; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachment to R⁵ H 0 0 1 —

TABLE 4 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q B —OCH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — C —HR¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 3 0 1 —

TABLE 5 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q G —CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 3 0 1 — D—CH₂OCH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 6 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q H —CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 1 1 1 — E —HR¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 1 1 1 —

TABLE 7 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q T —CH₃ R¹¹ is NR¹²; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ CH₃ 1 1 1 —

TABLE 8 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q AB —CH₂CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — Z —CH₃R¹¹ is S(O)_(q); and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 1 1 1 2

TABLE 9 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q ZZ —CH₃ R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 1 1 1 — O —CH₃R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attached taken togetherform:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 10 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q I —SCH(CH₃)₂ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹and R¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — J —CH₃R¹¹ is S(O)_(q); and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 0

TABLE 11 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹0, and R¹¹ R¹²n m p q P —CH₃ R¹¹ is S(O)_(q); and R⁹, R¹⁰, R¹¹, and the N to which R⁹and R¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachement to R⁵ — 0 0 1 2 Q —CH₃R¹¹ is CH₂; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachement to R⁵ — 1 1 1 —

TABLE 12 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q AC

R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — S

R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 13 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q V —CH₂OH R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachement to R⁵ — 0 0 1 — X —CH₃R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachement to R⁵ — 0 0 1 — EK—CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachement to R⁵ — 0 0 1 — EL—CH₃ R¹¹ is CF₂; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EM —CH₃R¹¹ is CF₂; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EN —CH₃R¹¹ is CF₂; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EO—CH₂OH R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EP—CH₂OH R¹¹ is CF₂; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — ER —CH₃R¹¹ is CF₂; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 14 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q AD

R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 15 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q K —SCH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 16 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹ R¹⁰ R¹¹ R¹² n m pq AF —CH₃ —H —CH₂CH₂OCH₃ — — 0 0 0 — AG —CH₃ —CH₃ —CH₂CH₂OCH₃ — — 0 0 0— M —CH₃ —CH₂CH₃ —CH₂CH₃ — — 0 0 0 — U —CH₃ —CH₂CH₃ —CH₂CH₃ — — 1 1 0 —ZY —CH₃ —H —CH₂CH₂OCH₃ — — 1 1 0 — AH —CH₃ —CH₂CH₃ —CH₂CH₃ — — 3 0 0 —AI —CH₃ —H —CH₂CH₂OCH₃ — — 3 0 0 — AJ —CH₃ —CH₃

— — 0 0 0 — ZX —CH₃ —H

— — 1 1 0 — Y —CH₃ —H

— — 1 1 0 —

TABLE 17 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹ R¹⁰ R¹¹ R¹² n m pq AK —CH₃ —H

— — 0 0 0 — W —CH₃ —H

— — 1 1 0 — KF —CH₃ —CH₃ —CH₃ — — 2 0 0 —

TABLE 18 Compounds having Formula I, wherein: R² = —CH₃, R³ =—CH₂CH(CH₃)₂, R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂O Compound R¹ R⁹ R¹⁰R¹¹ R¹² n m p q AL —CH₃ —CH₂CH₃ —CH₂CH₃ — — 2 0 0 —

TABLE 19 Compounds having Formula I, wherein: R² = —CH₃, R³ =—CH₂CH(CH₃)₂, R⁴ = —CH₃, R⁶ = —CH₂(CH₃)₂, and R⁸ = CH₂ Com- pound R¹ R⁹,R¹⁰, and R¹¹ R¹² n m p q AM —SCH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the Nto which R⁹ and R¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — AN —HR¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 20 Compounds having Formula I, wherein: R² = —CH₃, R³ =—CH₂CH(CH₃)₂, R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂ Com- pound R¹ R⁹,R¹⁰, and R¹¹ R¹² n m p q KG —CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the Nto which R⁹ and R¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 2 0 1 —

TABLE 21 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹ R¹⁰ R¹¹ R¹² n m pq DA —CH₃ —CH₃ —CH₂Pyrid-2-yl — — 0 0 0 — DB —CH₃ —CH₂Pyrid-2-yl—CH₂Pyrid-2-yl — — 0 0 0 — DC —CH₃ —CH₃ -Phenyl — — 0 0 0 — DD —CH₃ —CH₃-Pyrid-2-yl — — 0 0 0 — DE —CH₃ —CH₃ —CH₂CH₂SO₂NH₂ — — 0 0 0 — DF —CH₃—CH₃ —CH₂Pyrid-3-yl — — 0 0 0 — DG —CH₃ —CH₃ —CH₂Pyrimidin-2-yl — — 0 00 — DH —CH₃ —CH₃ —CH₂Pyrazin-2-yl — — 0 0 0 — DI —CH₃ —CH₃—CH₂-3-Me-Imidazol- — — 0 0 0 — 4-yl DJ —CH₃ —CH₃ —CH₂-2-Me-Pyrazol- — —0 0 0 — 3-yl DK —CH₃ —CH₃ —CH₂CH(CH₃)CN — — 0 0 0 — DL —CH₃ —CH₃—CH₂Pyrid-4-yl — — 0 0 0 — DM —CH₃ —CH₃ —CH₂-1-Me-Pyrazol- — — 0 0 0 —4-yl DN —CH₃ —CH₃ —CH₂CH₂CF₃ — — 0 0 0 — DO —CH₃ —CH₃ —CH₂-1-Me-3-CF₃- —— 0 0 0 — Pyrazol-5-yl DP —CH₃ —CH₃ —CH₂-5-F-Pyrid-2-yl — — 0 0 0 — DQ—CH₃ —CH₃ —CH₂-5-Cl-Pyrid-2-yl — — 0 0 0 — DR —CH₃ —CH₃—CH₂-3-CF₃-Pyrid-2- — — 0 0 0 — yl EQ —CH₃ —CH₂CH₂CF₃ —CH₂CH₂CF₃ — — 0 00 —

TABLE 22 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q DS —CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — DT —CH₃R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — DU —CH₃R¹¹ is N; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 23 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q DV —CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — DW —CH₃R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — DX —CH₃R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 24 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q DY —CH₃ R⁹, R¹⁰, R¹¹, and the N to whch R⁹ and R¹⁰ are attachedtaken together form:  

 wherein “*” represents the point of attachment to R⁵ — 0 0 1 — DZ —CH₃R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EA —CH₃R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 25 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q EB —CH₃ R¹¹ is CH₂; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EC —CH₃R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — ED —CH₃R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EE —CH₃R¹¹ is N; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EF —CH₃R¹¹ is CH₂; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 26 Compounds of Formula I, wherein: R² = —CH₃, R³ = —CH₂CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q EG —CH₃ R¹¹ is NR¹²; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ CH₃ 0 0 1 — EH—CH₃ R¹¹ is CH₂(OCH₃); and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 — EI —HR¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtaken together form:  

  wherein “*” represents the point of attachment to R⁵ — 1 0 1 —

TABLE 27 Compounds of Formula I, wherein: R² = —CH₂CH₃, R³ = —CH(CH₃)₂,R⁴ = —CH₃, R⁶ = —CH₂CH₃, and R⁸ = CH₂. Compound R¹ R⁹, R¹⁰, and R¹¹ R¹²n m p q EJ —CH₃ R¹¹ is O; and R⁹, R¹⁰, R¹¹, and the N to which R⁹ andR¹⁰ are attached taken together form:  

  wherein “*” represents the point of attachment to R⁵ — 0 0 1 —

TABLE 28 Cyclophilin A (Cyp A) Inhibitory Activity, immunosuppressivepotential, and aqueous solubility for select Compounds having Formula IProtease- Calcineurin Mixed free Phosphatase Lymphocyte Water PPIase(CaN) Reaction Solubility Assay* Assay (+CypA)** (MLR) Assay****Compound K_(i) (nM) IC₅₀ (nM) Assay*** (μM) Cyclo- 1.5 210 1 10-25 sporin A F 4.5 >10,000 144 >100 A 23 >10,000 >100 L 7.3 >10,000 541 >100LL 46 >10,000 N 6.1 >10,000 <100 NN 120 >10,000 B 8 >10,000 >100 G6.2 >10,000 77 >100 C 14 >10,000 84 >100 M 90 75-100 D 10 >10,000 >100075-100 H 3.9 >10,000 418 >100

TABLE 29 Data Showing Cyclophilin A (Cyp A) Inhibitory Activity,immunosuppressive potential, and aqueous solubility for select Compoundshaving Formula I Protease- Calcineurin Mixed free Phosphatase LymphocyteWater PPIase (CaN) Reaction Solubility Assay* Assay (+CypA)** (MLR)Assay**** Compound K_(i) (nM) IC₅₀ (nM) Assay*** (μM) T 5 >10,000 >100 U4 >10,000 75-100 ZY 5.7 >10,000 75-100 K 4.9 >10,000 >1000 >100 AB5.6 >10,000 >100 Y 4.1 >10,000 >1000 >100 AK 42 >10,000 W1.9 >10,000 >100 Z 2 >10,000 >100 ZZ 2.6 >10,000 >100 E 24 >10,000

TABLE 30 Data Showing Cyclophilin A (Cyp A) Inhibitory Activity,immunosuppressive potential, and aqueous solubility for select Compoundshaving Formula I Protease- Calcineurin Mixed free Phosphatase LymphocyteWater PPIase (CaN) Reaction Solubility Assay* Assay (+CypA)** (MLR)Assay**** Compound K_(i) (nM) IC₅₀ (nM) Assay*** (μM) O8.2 >10,000 >1000 >100 AF 28 >100 AG 18 >100 I 7.7 >10,000 >1000 25-50AH 4.7 >10,000 >100

TABLE 31 Data Showing Cyclophilin A (Cyp A) Inhibitory Activity,immunosuppressive potential, and aqueous solubility for select Compoundshaving Formula I Protease- Calcineurin Mixed free Phosphatase LymphocyteWater PPIase (CaN) Reaction Solubility Assay* Assay (+CypA)** (MLR)Assay**** Compound K_(i) (nM) IC₅₀ (nM) Assay*** (μM) AI3.3 >10,000 >100 AJ 12 50-75 J 3.7 6,300 50-75 P 3.2 >10,000 >100 Q3.5 >10,000 50-75 ZX 2.3 >10,000 >100

TABLE 32 Data Showing Cyclophilin A (Cyp A) Inhibitory Activity,immunosuppressive potential, and aqueous solubility for select Compoundshaving Formula I Protease- Calcineurin Mixed free Phosphatase LymphocyteWater PPIase (CaN) Reaction Solubility Assay* Assay (+CypA)** (MLR)Assay**** Compound K_(i) (nM) IC₅₀ (nM) Assay*** (μM) AC7.5 >10,000 >100 S 10 >10,000 >100 V 4.5 >10,000 >100 X 4.9 >10,000 >100AD 18 >10,000 AL 27 >10,000 >100 AN 15 >10,000 AM 1.9 >8,900 25-50 KF7.8 >10,000 KG 3.8 >10,000

TABLE 33 Data Showing Cyclophilin A (Cyp A) Inhibitory Activity,immunosuppressive potential, and aqueous solubility for select Compoundshaving Formula I Protease- Calcineurin Mixed free Phosphatase LymphocyteWater PPIase (CaN) Reaction Solubility Assay* Assay (+CypA)** (MLR)Assay**** Compound K_(i) (nM) IC₅₀ (nM) Assay*** (μM) DA 4.9 >10,000 75-100 DB 7.4 >10,000  75-100 DC 6.0 >10,000 10-25 DD 4.8 >10,000 25-50DE 5.8 >10,000 >100 DF 14 >10,000 >100 DG 8.5 >10,000 >100 DH9.1 >10,000 >100 DI 4.3 >10,000 >100 DJ 5.4 >10,000 >100 DK 136,300 >100 DL 5.7 >10,000 >100 DM 23 DN 5.5 >10,000 25-50 DO 2.8 >10,00010-25 DP 16.2 >10,000 DQ 11 >10,000 DR 8.4 >10,000

TABLE 34 Data Showing Cyclophilin A (Cyp A) Inhibitory Activity,immunosuppressive potential, and aqueous solubility for select Compoundshaving Formula I Protease- Calcineurin Mixed free Phosphatase LymphocyteWater PPIase (CaN) Reaction Solubility Assay* Assay (+CypA)** (MLR)Assay**** Compound K_(i) (nM) IC₅₀ (nM) Assay*** (μM) DS 12 >10,000 75-100 DT 4.7 >10,000 >100 DU 9.2 >10,000 >100 DV 4.3 >10,000 250 50-75DW 6.5 >10,000 >100 DX 5.2 7,600 250 25-50 DY 13 >10,000 >100 DZ6.0 >10,000 25-50 EA 22 >10,000 EF 40 >10,000 EB 3.7 >10,000 EC5.2 >10,000 ED 3.8 >10,000 EE 9.1 >10,000

TABLE 35 Data Showing Cyclophilin A (Cyp A) Inhibitory Activity,immunosuppressive potential, and aqueous solubility for select Compoundshaving Formula I Protease- Calcineurin Mixed free Phosphatase LymphocyteWater PPIase (CaN) Reaction Solubility Assay* Assay (+CypA)** (MLR)Assay**** Compound Ki (nM) IC50 (nM) Assay*** (μM) EJ 9.0 >10,000 >100EG 8.2 >10,000 >100 EH 17 >10,000 >100 EI 14 >10,000 >100 EK 12 225 1025-50 EL 10 1,300 10  75-100 EM 9.5 655 10  75-100 EN 6.7 2,400 50 25-50EQ 4.6 780 10 25-50 ER 6.7 1,600 25-50 EO 13 1,500 50 10-25 EP 3.4 7,80050-75

In Tables 28-35:

* Data generated using the protease-free PPIase assay.** Data generated using the Calcineurin Phosphatase (CaN) Assay. Nosignificant inhibition of CaN was observed in the absence or presence ofCypA. Data obtained in the presence of Cyp A (+CypA) are reported in thetable.*** Data generated using the Mixed Lymphocyte Reaction (“MLR”) Assay.The values shown are expressed as the IC50 for the compound relative tothe IC50 for Cyclosporin A. Thus, a value of 10, for example, indicatesthat the compound is about ten times less immunosuppressive thanCyclosporin A.**** Data generated using the Water Solubility Assay.

1. A compound having Formula I

or a pharmaceutically acceptable salt thereof, wherein: R¹ is —H,—C₁₋₆alkyl, —OC₁₋₆alkyl, —CH₂F, —CH₂OCH₃, —SC₁₋₆alkyl, —CH₃, —CH₂CH₃,—SCH(CH₃)₂, —CH₂OH, —SCH₃, —OCH₃, —R¹³R¹⁴,

R² is —CH₃, —CH₂CH₃, or —CH₂CH₂CH₃; R³ is —CH₂CH(CH₃)₂, —CH(CH₃)₂,—CH₂C(CH₃)₂(OH), —CH(CH₃)(CH₂CH₃), or —CH₂CH(R⁷)(CH₂CH₃); R⁴ is —CH₃ or—CH₂OH; R⁵ is —R⁸(CH₂)_(n)(C═O)_(m)—; R⁶ is —CH₂CH₃, —CH(CH₃)(OH),—CH(CH₃)₂, or —CH₂CH₂CH₃; R⁷ is OC₁₋₅ alkyl; R⁸ is O, S, CH₂O, CH₂S, orCH₂; R⁹ is —H, —C₁₋₅alkyl, —C₂₋₄fluoroalkyl, C₁₋₆alkyl-heterocycle,cyanoalkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or R⁹ taken togetherwith R¹¹, R¹⁰, and the N to which R⁹ and R¹⁰ are attached forms aheterocycle; R¹⁰ is —H, —C₁₋₅alkyl, —C₂₋₄fluoroalkyl,C₁₋₆alkyl-heterocycle, cyanoalkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or R¹⁰ takentogether with R¹¹, R⁹, and the N to which R⁹ and R¹⁰ are attached formsa heterocycle; R¹¹ is O, NR¹², S(O)_(q), CF₂, CH(OC₁₋₆alkyl),C₁₋₅alkylene, divalent C₃₋₈cycloalkyl, divalent heterocycle, carbonyl,or taken together with R⁹, R¹⁰, and the N to which R⁹ and R¹⁰ areattached forms a heterocycle; R¹² is H, CH₃, or C₁₋₅ alkyl; R¹³ is O, S,CH₂O, CH₂S, CH₂SO, or CH₂SO₂; R¹⁴ is H, C₁₋₆alkyl, C₂₋₆alkenyl,C₂₋₆alkynyl, C₃₋₈cycloalkyl, CH₂CH₂N(CH₂CH₃)₂, CH₂CH₂NH(CH₂CH₃),heterocycle, or aryl; n=0, 1, 2, 3, or 4; m=0 or 1; p=0 or 1; and q=0,1, or 2; wherein R¹⁴ is optionally substituted with one or more groupsindependently selected from the group consisting of H, C₁₋₆alkyl,halogen, hydroxyl, ester, sulfonamide, ketone, aldehyde, cycloalkyl,heterocycle, aryl, amine, heterocycle, amide, and guanidinyl; whereinthe heterocycle comprising R⁹, R¹⁰, R¹¹ and the N to which R⁹ and R¹⁰are attached is monocyclic or polycyclic; wherein “- - -” is a singlebond or is absent; and with the provisos that when R⁸ is O, S, CH₂O, orCH₂S then n is not 0 or 1; when p=0 then R¹¹ and “- - -” are absent; andwhen R¹¹ and “- - -” are absent then R⁹ is not directly linked to R¹⁰.2. A compound according to claim 1, or a pharmaceutically acceptablesalt thereof, wherein: R¹ is —H, —C₁₋₆alkyl, —OC₁₋₆alkyl, —CH₂F,—CH₂OCH₃, —SC₁₋₆alkyl, —CH₃, —CH₂CH₃, —SCH(CH₃)₂, —CH₂OH, —SCH₃, —OCH₃,—R¹³R¹⁴,

R² is —CH₃, —CH₂CH₃, or —CH₂CH₂CH₃; R³ is —CH₂CH(CH₃)₂, —CH(CH₃)₂,—CH₂C(CH₃)₂(OH), —CH(CH₃)(CH₂CH₃), or —CH₂CH(R⁷)(CH₂CH₃); R⁴ is —CH₃, or—CH₂OH; R⁵ is —R⁸(CH₂)_(n)(C═O)_(m)— R⁶ is —CH₂CH₃, —CH(CH₃)(OH),—CH(CH₃)₂, or —CH₂CH₂CH₃; R⁷ is OC₁₋₅ alkyl; R⁸ is O, S, CH₂O, CH₂S, orCH₂; R⁹ is —H, —C₁₋₅alkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or taken togetherwith R¹¹, R¹⁰, and the N to which R⁹ and R¹⁰ are attached forms aheterocycle; R¹⁰ is —H, —C₁₋₅alkyl,

C₃₋₈cycloalkyl, heterocycle, aryl, or cycloalkenyl, or taken togetherwith R¹¹, R⁹, and the N to which R⁹ and R¹⁰ are attached forms aheterocycle; R¹¹ is O, NR¹², S(O)_(q), C₁₋₅alkylene, divalentC₃₋₈cycloalkyl, divalent heterocycle, carbonyl, or taken together withR⁹, R¹⁰, and the N to which R⁹ and R¹⁰ are attached forms a heterocycle;R¹² is H, CH₃, or C₁₋₅ alkyl; R¹³ is O, S, CH₂O, CH₂S, CH₂SO, or CH₂SO₂;R¹⁴ is H, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, C₃₋₈cycloalkyl,CH₂CH₂N(CH₂CH₃)₂, CH₂CH₂NH(CH₂CH₃), heterocycle, or aryl; n=0, 1, 2, 3,or 4; m=0 or 1; p=0 or 1; and q=0, 1, or 2; wherein R¹⁴ is optionallysubstituted with one or more groups independently selected from thegroup consisting of H, C₁₋₆alkyl, halogen, hydroxyl, ester, sulfonamide,ketone, aldehyde, cycloalkyl, heterocycle, aryl, amine, heterocycle,amide, and guanidinyl; wherein the heterocycle comprising R⁹, R¹⁰, R¹¹,and the N to which R⁹ and R¹⁰ are attached is monocyclic or polycyclic;wherein “- - -” is a single bond or is absent; and with the provisosthat when R⁸ is O, S, CH₂O, or CH₂S then n is not 0 or 1; when p=0 thenR¹¹ and “- - -” are absent; and when R¹¹ and “- - -” are absent then R⁹is not directly linked to R¹⁰.
 3. A compound according to claim 1, withthe further proviso that when R² is —CH₃, and R³ is —CH₂CH(CH₃)₂, and R⁴is —CH₃, and R⁶ is —CH₂CH₃, then the group

is not —CH₂CH₂CH₂(C═O)N(CH₂CH₃)₂, —CH₂CH₂CH₂(C═O)N(CH₃)₂,—CH₂CH₂CH₂(C═O)NH₂, —CH₂CH₂CH₂CH₂CH₂NH(C═O)CH₃, —CH₂CH₂CH₂CH₂NH(C═O)CH₃,—CH₂CH₂CH₂CH₂CH₂NH(C═O)CH₂CH₃, —CH₂CH₂CH₂CH₂CH₂NH(C═O)CH₂CH₂ CH₃,—CH₂CH₂CH₂CH₂NH(C═O)CH₃, —CH₂NHCH₂COOH, or —CH₂NH(CH₂)₅COOH.
 4. Acompound according to claim 1 with the further provisos that when m=1,and R² is —CH₃, and R³ is —CH₂CH(CH₃)₂, and R⁴ is —CH₃, and R⁶ is—CH₂CH₃ then neither R⁹ nor R¹⁰ is —H or —C₁-C₅alkyl, and R⁹, R¹⁰, R¹¹,and the N to which R⁹ and R¹⁰ are attached taken together do not formmorpholinyl.
 5. A compound according to claim 1, wherein m=0 and R¹ isnot —H.
 6. A compound according to claim 2, wherein n=0, 1, 2, 3, or 4and m=0.
 7. A compound according to claim 2, wherein n=0 or 3, m=0, andp=1.
 8. A compound according to claim 2, wherein R¹ is not hydrogen (H).9. (canceled)
 10. (canceled)
 11. (canceled)
 12. (canceled)
 13. Acompound according to claim 2, wherein R² is —CH₃, R³ is —CH₂CH(CH₃)₂,R⁴ is —CH₃, R⁶ is —CH₂CH₃, and R⁸ is CH₂.
 14. (canceled)
 15. (canceled)16. (canceled)
 17. (canceled)
 18. (canceled)
 19. A compound according toclaim 2, wherein R¹ is —CH₃, R⁶ is —CH₂CH₃ or —CH(CH₃)₂, R⁸ is CH₂, R¹¹is O, n=0, 1, 2, 3, or 4, p=1, and R⁹, R¹⁰, R¹¹, and the N to which R⁹and R¹⁰ are attached taken together form morpholinyl.
 20. (canceled) 21.(canceled)
 22. A compound according to claim 2, wherein R¹═—CH₃,R²═—CH₃, R³=—CH₂CH(CH₃)₂, R⁴═—CH₃, R⁶═—CH₂CH₃, R⁸═CH₂, R¹¹ is O, n=0 or3, m=0, p=1, and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ areattached taken together form:

wherein “*” represents the point of attachment to R⁵.
 23. A compoundaccording to claim 1, wherein m=0, n=0, 1, 2, 3, or 4, and p=0 or
 1. 24.(canceled)
 25. (canceled)
 26. A compound according to claim 23, whereinp=1 and R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰ are attachedtogether form a heterocycle.
 27. A compound according to claim 26,wherein the heterocycle formed from R⁹, R¹⁰, R¹¹ and the N to which R⁹and R¹⁰ are attached is selected from the group consisting ofmorpholinyl, piperazinonyl, N-methylpiperazinyl,

wherein “*” represents the point of attachment to R⁵.
 28. A compoundaccording to claim 1, wherein R¹ is —CH₃, —SCH₃, —OCH₃, or —CH₂OH; R⁸ isCH₂; n=0, 1, 2, 3, or 4; m=0; p=1, and R⁹, R¹⁰, R¹¹, and the N to whichR⁹ and R¹⁰ are attached taken together form a heterocycle. 29.(canceled)
 30. (canceled)
 31. A compound according to claim 28, whereinthe heterocycle formed by R⁹, R¹⁰, R¹¹, and the N to which R⁹ and R¹⁰are attached taken together is substituted by an alkyl, halogen, orhaloalkyl.
 32. A compound according to claim 31, wherein saidsubstituted heterocycle is substituted by —CF₃ or fluorine.
 33. Acompound according to claim 1 selected from the group consisting of:[(3R,4R,5S)-4-(hydroxy)-3-methyl-5-(methylamino)-1-(morpholin-4-yl)hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Compound F);[(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-(4-methylpiperazino)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Compound L);[(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-diethylamino-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Compound M);[(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-(N-3-piperazinone)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Compound N);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-{1H-imidazol-4-yl}-ethylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AK);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-methoxyethylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AF);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-methoxyethyl)methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AG);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(N-(3aR*,6aS*)-2-methyloctahydropyrrolo[3,4-c]pyrrolo)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound O);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(1,4-dioxan-2-ylmethyl)methylamino-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AJ);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(thiomorpholino)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Compound J);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(1,1-dioxo-thiomorpholino)-hexanoicacid]¹[(R)-Me-Sar]³cyclosporin A (Compound P);[[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-homomorpholino-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound X);[(6R,7R,8S)-7-hydroxy-6-methyl-8-(methylamino)-1-N-morpholino-nonanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound G);[(6R,7R,8S)-7-Hydroxy-6-methyl-8-(methylamino)-1-N-diethylamino-nonanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AH);[(6R,7R,8S)-7-Hydroxy-6-methyl-8-(methylamino)-1-N-(2-methoxy)ethylamino-nonanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound AI);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-N-morpholino-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound H);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(4-methylpiperazin-1-yl)-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound T);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-diethylamino-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound U);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-{sulfonic aciddimethylamide}-ethylamino)-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound W);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-{1H-imidazol-4-yl}-ethylamino)-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound Y);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-({1,1-dioxo}thiomorpholin-4-yl)-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound Z);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(N-(3aR*,6aS*)-2-methyloctahydropyrrolo[3,4-c]pyrrolo)-1-oxo-octanoicacid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZZ);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(2-methoxyethylamino)-1-oxo-octanoicacid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZY);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(1,4-dioxan-2-ylmethyl)amino)-1-oxo-octanoicacid]1[(R)-methyl-Sar]3cyclosporin A (Compound ZX);[(5R,6R,7S)-6-Hydroxy-5-methyl-7-(methylamino)-1-piperidino-1-oxo-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound Q);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-2-(N,N-diethylamino)ethoxy-hexanoicacid]¹[(S)-thio-isopropyl-Sar]³cyclosporin A (Compound AL);[(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-ethyl-Sar]³cyclosporin A (Compound AB);[(3R,4R,5S)-4-hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-thiomethyl-Sar]³cyclosporin A (Compound K);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-thio-isopropyl-Sar]³cyclosporin A (Compound I);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-methoxy-Sar]³cyclosporin A (Compound B);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-methoxymethylene-Sar]³cyclosporin A (Compound D);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-hydroxymethyl-Sar]³cyclosporin A (Compound V);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(R)-2 diethylamino ethyl oxymethyl-Sar]³cyclosporin A (CompoundS);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-homomorpholino-hexanoicacid]¹[(R)-2 diethylamino ethyl oxymethyl-Sar]³cyclosporin A (CompoundAD);[(5R,6R,7S)-1-(dimethylamino)-6-hydroxy-5-methyl-7-(methylamino)-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound KF);[(5R,6R,7S)-6-hydroxy-5-methyl-7-(methylamino)-1-(N-morpholino)-octanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound KG);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-N-morpholino-hexanoicacid]¹[(S)-2-diethylaminoethylthiomethyl-Sar]³cyclosporinA (CompoundAC);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DA);[(3R,4R,5S)-1-(Bis{pyridin-2-ylmethyl}amino)-4-hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DB);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(methyl-phenyl-amino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DC);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(methyl-pyridin-2-yl-amino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DD);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-sulfamoyl-ethyl)-methyl-amino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DE);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-3-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DF);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyrimidin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DG);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyrazin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DH);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3-methyl-3H-imidazol-4-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DI);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({2-methyl-2H-pyrazol-3-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DJ);[(3R,4R,5S)-1-({2-Cyano-propyl}-methyl-amino)-4-hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DK);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({pyridin-4-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DL);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({1-methyl-1H-pyrazol-4-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DM);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3,3,3-trifluoropropyl}-methyl-amino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DN);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({1-methyl-3-trifluoromethyl-2H-pyrazol-5-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DO);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({5-fluoro-pyridin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DP);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({5-chloro-pyridin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DQ);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-({3-trifluoromethyl-pyridin-2-ylmethyl}-methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DR);[(3R,4R,5S)-1-(3,3-Dimethyl-morpholin-4-yl)-4-hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DS);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-methylamino-((R)-3-methyl-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DT);[(3R,4R,5S)-1-(5,6-Dihydro-8H-[1,2,4]triazolo[4,3-a]pyrazin-7-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DU);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(8-oxa-3-aza-bicyclo[3.2.1]oct-3-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DV);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-((S)-3-methyl-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DW);[(3R,4R,5S)-1-(2,3-Dihydro-benzo[1,4]oxazin-4-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DX);[(3R,4R,5S)-1-(1,4-Dioxa-8-aza-spiro[4.5]dec-8-yl)-4-Hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DY);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-phenyl-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound DZ);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(piperidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EA);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(pyrrolidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EF);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-trifluoromethyl-piperidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EB);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(3-trifluoromethyl-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EC);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-([1,2]oxazinan-2-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound ED);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2-trifluoromethyl-5,6-dihydro-8H-imidazo[1,2-a]pyrazin-7-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EE);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(pyrrolidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EF);[[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(4-methyl-[1,4]diazepan-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EG);[[(3R,4R,5S)-4-Hydroxy-1-(3-methoxy-azetidin-1-yl)-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EH);[(4R,5R,6S)-5-Hydroxy-4-methyl-6-(methylamino)-1-(morpholin-4-yl)-heptanoicacid]¹cyclosporin A (Compound EI);[(3R,4R,5S)-4-Hydroxy-3-methyl-1-(morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³[Ethyl-Val]⁴cyclosporin A (Compound EJ);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2,2,6,6-tetrafluoro-morpholin-4-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EK);[(3R,4R,5S)-(3,3-Difluoro-pyrrolidin-1-yl)-4-Hydroxy-3-methyl-5-(methylamino)-1-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EL);[(3R,4R,5S)-(3,3-Difluoro-azetidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-1-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EM);[(3R,4R,5S)-(4,4-Difluoro-piperidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-1-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EN);[(3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(3,3,4,4-tetrafluoro-pyrrolidin-1-yl)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound ER);3R,4R,5S)-4-Hydroxy-3-methyl-5-(methylamino)-1-(2,2,6,6-tetrafluoro-morpholin-4-yl)-hexanoicacid]¹[(R)-hydroxymethyl-Sar]³cyclosporin A (Compound EO);(3R,4R,5S)-1-(3,3-Difluoro-pyrrolidin-1-yl)-4-hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-hydroxymethyl-Sar]³cyclosporin A (Compound EP); and(3R,4R,5S)-1-[Bis-(3,3,3-trifluoro-propyl)-amino]-[(4-Hydroxy-3-methyl-5-(methylamino)-hexanoicacid]¹[(R)-methyl-Sar]³cyclosporin A (Compound EQ).
 34. A pharmaceuticalcomposition comprising as active ingredient a therapeutically effectiveamount of a compound according to claim 1, or a pharmaceuticallyacceptable salt thereof, and a pharmaceutically acceptable excipient.35. (canceled)
 36. The pharmaceutical composition of claim 34, whereinthe composition is in the form of an emulsion or an aqueous solution.37. The pharmaceutical composition of claim 34, wherein the compositionis acceptable for administration to the eye(s) of a mammal.
 38. A methodof treating a medical condition in a patient in need thereof, the methodcomprising administering to the patient a therapeutically effectiveamount of a compound according to claim 1, or a pharmaceuticallyacceptable salt thereof.
 39. (canceled)
 40. (canceled)
 41. (canceled)42. (canceled)
 43. A method for treating dry eye in a patient in needthereof, the method comprising administering to the eye(s) of thepatient a therapeutically effective amount of a compound, or apharmaceutically acceptable salt thereof, according to claim
 1. 44. Amethod for increasing tear production in a patient whose tear productionis suppressed or presumed to be suppressed due to ocular inflammationassociated with keratoconjunctivitis sicca, the method comprisingadministering to the eye(s) of the patient a therapeutically effectiveamount of a compound according to claim 1, or a pharmaceuticallyacceptable salt thereof.
 45. (canceled)
 46. A method for reducing orpreventing an ocular condition in a patient, the method comprisingadministering a therapeutically effective amount of a compound accordingto claim 1 to the patient, wherein the ocular condition is selected fromthe group consisting of dry eye, ocular surface inflammation,blepharitis, meibomian gland disease, allergic conjunctivitis, pterygia,ocular symptoms of graft versus host disease, ocular allergy, atopickeratoconjunctivitis, vernal keratoconjunctivitis, uveitis, anterioruveitis, Behcet's disease, ocular cicatricial pemphigoid, chronic ocularsurface inflammation caused by viral infection, herpes simplexkeratitis, adenoviral keratoconjunctivitis, ocular rosacea, andpinguecula. 47.-49. (canceled)